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  • 1.
    Alam, Md Khorshed
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Vinklarek, Ivo
    Gröbner, Gerhard
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Johansson, Lennart B-Å
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Sachl, Radek
    Fluorescence Studies of Lipid Distribution in Bilayers under Oxidative Stress2019Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 116, nr 3, s. 508A-508AArtikkel i tidsskrift (Annet vitenskapelig)
  • 2.
    Andersson, Magnus
    et al.
    Umeå universitet, Teknisk-naturvetenskaplig fakultet, Fysik.
    Fällman, Erik
    Umeå universitet, Teknisk-naturvetenskaplig fakultet, Fysik.
    Uhlin, Bernt Eric
    Umeå universitet, Medicinsk fakultet, Molekylärbiologi (Medicinska fakulteten).
    Axner, Ove
    Umeå universitet, Teknisk-naturvetenskaplig fakultet, Fysik.
    A sticky chain model of the elongation and unfolding of escherichia coli P pili under stress2006Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 90, nr 5, s. 1521-1534Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    A model of the elongation of P pili expressed by uropathogenic Escherichia coli exposed to stress is presented. The model is based upon the sticky chain concept, which is based upon Hooke’s law for elongation of the layer-to-layer and head-to-tail bonds between neighboring units in the PapA rod and a kinetic description of the opening and closing of bonds, described by rate equations and an energy landscape model. It provides an accurate description of the elongation behavior of P pili under stress and supports a hypothesis that the PapA rod shows all three basic stereotypes of elongation/unfolding: elongation of bonds in parallel, the zipper mode of unfolding, and elongation and unfolding of bonds in series. The two first elongation regions are dominated by a cooperative bond opening, in which each bond is influenced by its neighbor, whereas the third region can be described by individual bond opening, in which the bonds open and close randomly. A methodology for a swift extraction of model parameters from force-versus-elongation measurements performed under equilibrium conditions is derived. Entities such as the free energy, the stiffness, the elastic elongation, the opening length of the various bonds, and the number of PapA units in the rod are determined.

  • 3.
    Andersson, Magnus
    et al.
    Umeå universitet, Teknisk-naturvetenskaplig fakultet, Fysik.
    Fällman, Erik
    Umeå universitet, Teknisk-naturvetenskaplig fakultet, Fysik.
    Uhlin, Bernt Eric
    Umeå universitet, Medicinsk fakultet, Molekylärbiologi (Medicinska fakulteten).
    Axner, Ove
    Umeå universitet, Teknisk-naturvetenskaplig fakultet, Fysik.
    Dynamic Force Spectroscopy of E. coli P Pili2006Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 91, nr 7, s. 2717-2725Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Surface organelles (so-called pili) expressed on the bacterial membrane mediate the adhesion of Escherichia coli causing urinary tract infection. These pili possess some extraordinary elongation properties that are assumed to allow a close bacterium-to-host contact even in the presence of shear forces caused by urine flow. The elongation properties of P pili have therefore been assessed for low elongation speeds (steady-state conditions). This work reports on the behavior of P pili probed by dynamic force spectroscopy. A kinetic model for the unfolding of a helixlike chain structure is derived and verified. It is shown that the unfolding of the quaternary structure of the PapA rod takes place at a constant force that is almost independent of elongation speed for slow elongations (up to ~0.4 μm/s), whereas it shows a dynamic response with a logarithmic dependence for fast elongations. The results provide information about the energy landscape and reaction rates. The bond length and thermal bond opening and closure rates for the layer-to-layer bond have been assessed to ~0.76 nm, ~0.8 Hz, and ~8 GHz, respectively. The results also support a previously constructed sticky-chain model for elongation of the PapA rod that until now had been experimentally verified only under steady-state conditions.

  • 4.
    Andersson, Magnus
    et al.
    Umeå universitet, Teknisk-naturvetenskaplig fakultet, Fysik.
    Uhlin, Bernt Eric
    Umeå universitet, Medicinsk fakultet, Molekylärbiologi (Medicinska fakulteten).
    Fällman, Erik
    Umeå universitet, Teknisk-naturvetenskaplig fakultet, Fysik.
    The biomechanical properties of E. coli pili for urinary tract attachment reflect the host environment2007Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 93, nr 9, s. 3008-3014Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Uropathogenic Escherichia coli express pili that mediate binding to host tissue cells. We demonstrate with in situ force measuring optical tweezers that the ability of P and type 1 pili to elongate by unfolding under exposure to stress is a shared property with some differences. The unfolding force of the quaternary structures under equilibrium conditions is similar, 28 ± 2 and 30 ± 2 pN for P pili and type 1 pili, respectively. However, type 1 pili are found to be more rigid than P pili through their stronger layer-to-layer bonds. It was found that type 1 pili enter a dynamic regime at elongation speeds of 6 nm/s, compared to 400 nm/s for P pili; i.e., it responds faster to an external force. This possibly helps type 1 to withstand the irregular urine flow in the urethra as compared to the more constant urine flow in the upper urinary tract. Also, it was found that type 1 pili refold during retraction at two different levels that possibly could be related to several possible configurations. Our findings highlight functions that are believed to be of importance for the bacterial ability to sustain a basic antimicrobial mechanism of the host and for bacterial colonization.

  • 5.
    Björnham, Oscar
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för fysik.
    Axner, Ove
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för fysik.
    Catch-Bond behavior of bacteria binding by slip bonds2010Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 99, nr 5, s. 1331-1341Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    It is shown that multipili-adhering bacteria expressing helix-like pili binding by slip bonds can show catch-bond behavior. When exposed to an external force, such bacteria can mediate adhesion to their hosts by either of two limiting means: sequential or simultaneous pili force exposure (referring to when the pili mediate force in a sequential or simultaneous manner, respectively). As the force is increased, the pili can transition from sequential to simultaneous pili force exposure. Since the latter mode of adhesion gives rise to a significantly longer bacterial adhesion lifetime than the former, this results in a prolongation of the lifetime, which shows up as a catch-bond behavior. The properties and conditions of this effect were theoretically investigated and assessed in some detail for dual-pili-adhering bacteria, by both analytical means and simulations. The results indicate that the adhesion lifetime of such bacteria can be prolonged by more than an order of magnitude. This implies that the adhesion properties of multibinding systems cannot be directly conveyed to the individual adhesion-receptor bonds.

  • 6. Chamberlain, Aaron K
    et al.
    MacPhee, Cait E
    Zurdo, Jesús
    Morozova-Roche, Ludmilla A
    Oxford Centre for Molecular Sciences, New Chemistry Laboratory, University of Oxford.
    Hill, H Allen O
    Dobson, Christopher M
    Davis, Jason J
    Ultrastructural organization of amyloid fibrils by atomic force microscopy2000Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 79, nr 6, s. 3282-3293Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Atomic force microscopy has been employed to investigate the structural organization of amyloid fibrils produced in vitro from three very different polypeptide sequences. The systems investigated are a 10-residue peptide derived from the sequence of transthyretin, the 90-residue SH3 domain of bovine phosphatidylinositol-3'-kinase, and human wild-type lysozyme, a 130-residue protein containing four disulfide bridges. The results demonstrate distinct similarities between the structures formed by the different classes of fibrils despite the contrasting nature of the polypeptide species involved. SH3 and lysozyme fibrils consist typically of four protofilaments, exhibiting a left-handed twist along the fibril axis. The substructure of TTR(10-19) fibrils is not resolved by atomic force microscopy and their uniform appearance is suggestive of a regular self-association of very thin filaments. We propose that the exact number and orientation of protofilaments within amyloid fibrils is dictated by packing of the regions of the polypeptide chains that are not directly involved in formation of the cross-beta core of the fibrils. The results obtained for these proteins, none of which is directly associated with any human disease, are closely similar to those of disease-related amyloid fibrils, supporting the concept that amyloid is a generic structure of polypeptide chains. The detailed architecture of an individual fibril, however, depends on the manner in which the protofilaments assemble into the fibrillar structure, which in turn is dependent on the sequence of the polypeptide and the conditions under which the fibril is formed.

  • 7.
    Christiansen, Alexander
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Wittung-Stafshede, Pernilla
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Quantification of excluded volume effects on the folding landscape of Pseudomonas aeruginosa Apoazurin In Vitro2013Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 105, nr 7, s. 1689-1699Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Proteins fold and function inside cells that are crowded with macromolecules. Here, we address the role of the resulting excluded volume effects by in vitro spectroscopic studies of Pseudomonas aeruginosa apoazurin stability (thermal and chemical perturbations) and folding kinetics (chemical perturbation) as a function of increasing levels of crowding agents dextran (sizes 20, 40, and 70 kDa) and Ficoll 70. We find that excluded volume theory derived by Minton quantitatively captures the experimental effects when crowding agents are modeled as arrays of rods. This finding demonstrates that synthetic crowding agents are useful for studies of excluded volume effects. Moreover, thermal and chemical perturbations result in free energy effects by the presence of crowding agents that are identical, which shows that the unfolded state is energetically the same regardless of method of unfolding. This also underscores the two-state approximation for apoazurin’s unfolding reaction and suggests that thermal and chemical unfolding experiments can be used in an interchangeable way. Finally, we observe increased folding speed and invariant unfolding speed for apoazurin in the presence of macromolecular crowding agents, a result that points to unfolded-state perturbations. Although the absolute magnitude of excluded volume effects on apoazurin is only on the order of 1–3 kJ/mol, differences of this scale may be biologically significant.

  • 8.
    Dingeldein, Artur P G
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Lidman, Martin
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Pokorna, Sarka
    Hof, Martin
    Pedersen, Anders
    Karlsson, Göran
    Gröbner, Gerhard
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    BCL-2 Family Proteins Effect on Mitochondrial-Mimicking Membrane Structure by Solid State NMR2015Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 108, nr 2, s. 251A-252AArtikkel i tidsskrift (Annet vitenskapelig)
    Abstract [en]

    Mitochondria are not only the cells' powerhouse, but also involved in their suicide via apoptosis. Key regulators of this pathway are members of the Bcl-2 protein family which interact with the outer mitochondrial membrane to modulate permeability and enable the release of apoptotic stimuli like cytochrome c. For a long time the mitochondrial membrane forming lipids have been seen as merely structural building units with proteins doing the actual work. This view changed in recent years, since lipids were shown to be also directly involved in apoptotic events e.g. under intracellular oxidative stress. Oxidized phospholipids (OxPls) generated under these stress conditions might trigger mitochondria-mediated apoptosis. Their presence in mitochondrial membranes can severely alter the properties of these membranes with yet unknown consequences regarding the formation of pores through membrane-mediated interplay with apoptotic Bax protein. We therefore devised a model system that embodies oxidative stress conditions by incorporating OxPls into mitochondria mimicking model membranes composed of phosphatidylcholine (PC), phosphatidylethanolamine (PE) and cardiolipin (CL) to study the impact of OxPls on apoptotic Bax-membrane interactions. To obtain molecular insight into hydrophobic fatty acid regions of membranes and their hydrophilic interface which is responsible for first protein-membrane contacts, we used differential scanning calorimetry (DSC) and solid state NMR spectroscopy. Upon incorporating OxPls with carboxyl (PoxnoPC) or aldehyde (PazePC) groups at their truncated sn-2-chains into our mitochondria model membranes, calorimetric and NMR measurements showed dramatic changes. 31P NMR experiments revealed major perturbation effects in these membranes; an effect which presumably elevates the membrane binding of apoptotic Bax to the charged membranes and its partial penetration, being a prerequisite for its final formation of pores which enable cytochrome c release from the mitochondrial interior. Currently structural studies of various Bax-lipid assemblies are ongoing.

  • 9.
    Dingeldein, Artur P. G.
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Lidman, Martin
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Sparrman, Tobias
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Gröbner, Gerhard
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Dynamical and Structural Alterations withing Lipid-Protein Assemblies Control Apoptotic Pore Formation - A Solid State NMR Study2016Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 110, nr 3, s. 59A-60AArtikkel i tidsskrift (Annet vitenskapelig)
  • 10.
    Dingeldein, Artur P. G.
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Sparrman, Tobias
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Ådén, Jörgen
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Wacklin, Hanna P.
    Clifton, Luke A.
    Gröbner, Gerhard
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Mitochondrial Membrane Organization under Oxidative Stress: Insight by Solid-State NMR and Neutron Reflectometry2019Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 116, nr 3, s. 508A-508AArtikkel i tidsskrift (Annet vitenskapelig)
  • 11.
    Dingeldein, Artur Peter Günther
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Pokorna, Sarka
    Lidman, Martin
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Sparrman, Tobias
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Sachl, Radek
    Hof, Martin
    Gröbner, Gerhard
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Apoptotic Bax at Oxidatively Stressed Mitochondrial Membranes: Lipid Dynamics and Permeabilization2017Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 112, nr 10, s. 2147-2158Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Mitochondria are crucial compartments of eukaryotic cells because they function as the cellular power plant and play a central role in the early stages of programmed cell death (apoptosis). To avoid undesired cell death, this apoptotic pathway is tightly regulated by members of the Bcl-2 protein family, which interact on the external surface of the mitochondria, i.e., the mitochondrial outer membrane (MOM), and modulate its permeability to apoptotic factors, controlling their release into the cytosol. A growing body of evidence suggests that the MOM lipids play active roles in this permeabilization process. In particular, oxidized phospholipids (OxPls) formed under intracellular stress seem to directly induce apoptotic activity at the MOM. Here we show that the process of MOM pore formation is sensitive to the type of OxPls species that are generated. We created MOM-mimicking liposome systems, which resemble the cellular situation before apoptosis and upon triggering of oxidative stress conditions. These vesicles were studied using P-31 solid-state magic-angle-spinning nuclear magnetic resonance spectroscopy and differential scanning calorimetry, together with dye leakage assays. Direct polarization and cross-polarization nuclear magnetic resonance experiments enabled us to probe the heterogeneity of these membranes and their associated molecular dynamics. The addition of apoptotic Bax protein to OxPls-containing vesicles drastically changed the membranes' dynamic behavior, almost completely negating the previously observed effect of temperature on the lipids' molecular dynamics and inducing an ordering effect that led to more cooperative membrane melting. Our results support the hypothesis that the mitochondrion-specific lipid cardiolipin functions as a first contact site for Bax during its translocation to the MOM in the onset of apoptosis. In addition, dye leakage assays revealed that different OxPls species in the MOM-mimicking vesicles can have opposing effects on Bax pore formation.

  • 12.
    Dingeldein, Artur Peter Günther
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Sparrman, Tobias
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Gröbner, Gerhard
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Oxidatively stressed mitochondrial membranes: A molecular insight into their organization and function during apoptosisInngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Mitochondria are crucially involved in the removal of eukaryotic cells by theintrinsic pathway of programmed cell death (apoptosis). The mitochondrial outermembrane (MOM) is the platform where this pathway is regulated. Uponoxidative stress that triggers apoptotic action, the MOM undergoespermeabilization and releases cytochrome c, ultimately causing cell death. Notonly is this membrane perforation regulated by opposing members of the Bcl-2protein family meeting at the MOM but also is it regulated by the membrane itselfthat plays an active role. Upon oxidative damage, the membrane undergoessevere reorganization causing an imbalance towards cell death-causing apoptoticBcl-2 proteins. To understand the active role of MOM, we provided a detailedmolecular view of its structural and dynamic reorganization by solid-state13C MAS NMR (magic angle spinning nuclear magnetic resonance) accompaniedby calorimetric studies. By focusing on MOM-like vesicles doped with oxidizedlipid species, direct polarization 13C MAS NMR provided a quantitative overviewand identification of all lipid moieties across the membrane. 1H-13C crosspolarization and insensitive nuclei enhanced by polarization transfer MAS NMRgenerated a dynamic - mobile versus restricted – membrane profile. Evidently,oxidized phospholipids significantly perturb the structural membraneorganization and increase membrane dynamics. However, these perturbationsare not observed uniformly as the hydrophobic core is reflecting the melting ofthe lipid chains and the increase in lipid molecule disorder directly, whereas theinterface and headgroup region undergo complex changes in their dynamics,which are coupled to increased segmental disorder. These changes are potentiallycrucial in augmenting the pro-apoptotic action of proteins like Bax there.

  • 13. Grønberg, Christina
    et al.
    Sitsel, Oleg
    Lindahl, Erik
    Gourdon, Pontus
    Andersson, Magnus
    Theoretical Physics and Swedish e-Science Research Center, Science for Life Laboratory, KTH Royal Institute of Technology, Solna, Sweden.
    Membrane Anchoring and Ion-Entry Dynamics in P-type ATPase Copper Transport2016Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 111, nr 11, s. 2417-2429Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Cu+-specific P-type ATPase membrane protein transporters regulate cellular copper levels. The lack of crystal structures in Cu+-binding states has limited our understanding of how ion entry and binding are achieved. Here, we characterize the molecular basis of Cu+ entry using molecular-dynamics simulations, structural modeling, and in vitro and in vivo functional assays. Protein structural rearrangements resulting in the exposure of positive charges to bulk solvent rather than to lipid phosphates indicate a direct molecular role of the putative docking platform in Cu+ delivery. Mutational analyses and simulations in the presence and absence of Cu+ predict that the ion-entry path involves two ion-binding sites: one transient Met148-Cys382 site and one intramembranous site formed by trigonal coordination to Cys384, Asn689, and Met717. The results reconcile earlier biochemical and x-ray absorption data and provide a molecular understanding of ion entry in Cu+-transporting P-type ATPases.

  • 14.
    Gupta, Arun
    et al.
    Umeå universitet.
    Reinartz, Ines
    Spilotros, Alessandro
    Jonna, Venkateswara R.
    Umeå universitet.
    Hofer, Anders
    Umeå universitet.
    Svergun, Dmitri I.
    Schug, Alexander
    Wolf-Watz, Magnus
    Umeå universitet.
    Global Disordering in Stereo-Specific Protein Association2017Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 112, nr 3, s. 33A-33AArtikkel i tidsskrift (Fagfellevurdert)
  • 15. Hoernke, Maria
    et al.
    Larsson, Elin
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Mohan, Jagan
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Blomberg, Jeanette
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Westenhoff, Sebastian
    Lundmark, Richard
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Schwieger, Christian
    Structural Mechanism in a Membrane Remodelling ATP-ASE2016Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 110, nr 3, s. 578A-578AArtikkel i tidsskrift (Annet vitenskapelig)
  • 16. Homouz, Dirar M
    et al.
    Perham, Michael
    Samiotakis, Antonios
    Cheung, Margaret
    Wittung-Stafshede, Pernilla
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Crowded, cell-like environment induces shape changes in aspherical protein2009Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 96, nr 3, Supplement 1, s. 568a-Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Protein dynamics in cells may be different from that in dilute solutions in vitro since the environment in cells is highly concentrated with other macromolecules. This volume exclusion due to macromolecular crowding is predicted to affect both equilibrium and kinetic processes involving protein conformational changes. To quantify macromolecular crowding effects on protein folding mechanisms, here we have investigated the folding energy landscape of an α/β protein, apoflavodoxin, in the presence of inert macromolecular crowding agents using in silico and in vitro approaches. By coarse-grained molecular simulations and topology-based potential interactions, we probed the effects of increased volume fraction of crowding agents (fc) as well as of crowding agent geometry (sphere or spherocylinder) at high fc. Parallel kinetic folding experiments with purified Desulfovibro desulfuricans apoflavodoxin in vitro were performed in the presence of Ficoll (sphere) and Dextran (spherocylinder) synthetic crowding agents. In conclusion, we have identified in silico crowding conditions that best enhance protein stability and discovered that upon manipulation of the crowding conditions, folding routes experiencing topological frustrations can be either enhanced or relieved. The test-tube experiments confirmed that apoflavodoxin's time-resolved folding path is modulated by crowding agent geometry. We propose that macromolecular crowding effects may be a tool for manipulation of protein folding and function in living cells.

  • 17.
    Humpolickova, Jana
    et al.
    J. Heyrovský Institute of Physical Chemistry, Prague, Czech Republic.
    Stefl, Martin
    J. Heyrovský Institute of Physical Chemistry, Prague, Czech Republic.
    Sachl, Radek
    J. Heyrovský Institute of Physical Chemistry, Prague, Czech Republic.
    Cebecauer, Marek
    J. Heyrovský Institute of Physical Chemistry, Prague, Czech Republic.
    Machan, Radek
    J. Heyrovský Institute of Physical Chemistry, Prague, Czech Republic.
    Johansson, Lennart B-Å
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Hof, Martin
    J. Heyrovský Institute of Physical Chemistry, Prague, Czech Republic.
    Dynamics and size of crosslinking-induced lipid nanodomains in model membranes2012Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 102, nr 3, s. 294a-Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Changes of membrane organization upon crosslinking of its components trigger cellsignaling response to various exogenous factors. Crosslinking of raft gangliosides GM1with cholera toxin (CTxB) was demonstrated to cause microscopic phase separation inmodel membranes and the CTxB-GM1 complexes forming a minimal lipid raft unit aresubject of ongoing cell membrane research. Yet, those subdiffraction sized rafts havenever been described in terms of size and dynamics. By means of two-color z-scanfluorescence correlation spectroscopy, we show that the nano-sized domains are formedin model membranes at lower sphingomyelin content than needed for the large scalephase separation and that the CTxB-GM1 complexes are confined in the domains poorlystabilized with sphingomyelin. Fluorescence resonance energy transfer together withMonte Carlo modeling of the donor decay response reveal the domain radius ofapproximately 8 nm, which increases at higher sphingomyelin content. We observed twotypes of differently behaving domains, which suggests a dual role of the crosslinker: first,local transient condensation of the GM1 molecules compensating lack of sphingomyelinand second, coalescence of existing nanodomains ending in large scale phase separation.

  • 18.
    Jass, Jana
    et al.
    Department of Microbiology and Immunology, The Lawson Health Research Institute, University of Western Ontario, London, Ontario, N6A 4V2, Canada.
    Schedin, Staffan
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för tillämpad fysik och elektronik.
    Fällman, Erik
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för fysik.
    Olsson, J.
    Nilsson, U.
    Uhlin, Bernt Eric
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Axner, Ove
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för fysik.
    Physical properties of Escherichia coli P pili measured by optical tweezers2004Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Biophysical Journal, Vol. 87, nr 6, s. 4271-4283Artikkel i tidsskrift (Fagfellevurdert)
  • 19.
    Kahra, Dana
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Kovermann, Michael
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen. Chemistry Department, University of Konstanz, Konstanz, Germany.
    Wittung-Stafshede, Pernilla
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen. Department of Biology and Biological Engineering, Chalmers University of Technology, Gothenburg, Sweden.
    The C-Terminus of Human Copper Importer Ctr1 Acts as a Binding Site and Transfers Copper to Atox12016Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 110, nr 1, s. 95-102Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Uptake of copper (Cu) ions into human cells is mediated by the plasma membrane protein Ctr1 and is followed by Cu transfer to cytoplasmic Cu chaperones for delivery to Cu-dependent enzymes. The C-terminal cytoplasmic tail of Ctr1 is a 13-residue peptide harboring an HCH motif that is thought to interact with Cu. We here employ biophysical experiments under anaerobic conditions in peptide models of the Ctr1 C-terminus to deduce Cu-binding residues, Cu affinity, and the ability to release Cu to the cytoplasmic Cu chaperone Atox1. Based on NMR assignments and bicinchoninic acid competition experiments, we demonstrate that Cu interacts in a 1:1 stoichiometry with the HCH motif with an affinity, K-D, of similar to 10(-14) M. Removing either the Cys residue or the two His residues lowers the Cu-peptide affinity, but site specificity is retained. The C-terminal peptide and Atox1 do not interact in solution in the absence of Cu. However, as directly demonstrated at the residue level via NMR spectroscopy, Atox1 readily acquires Cu from the Cu-loaded peptide. We propose that Cu binding to the Ctr1 C-terminal tail regulates Cu transport into the cytoplasm such that the metal ion is only released to high-affinity Cu chaperones.

  • 20. Karolin, J
    et al.
    Fa, M
    Wilczynska, M
    Ny, Tor
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Johansson, L B
    Donor-donor energy migration for determining intramolecular distances in proteins: I. Application of a model to the latent plasminogen activator inhibitor-1 (PAI-1).1998Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 74, nr 1, s. 11-21Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    A new fluorescence spectroscopic method is presented for determining intramolecular and intermolecular distances in proteins and protein complexes, respectively. The method circumvents the general problem of achieving specific labeling with two different chromophoric molecules, as needed for the conventional donor-acceptor transfer experiments. For this, mutant forms of proteins that contain one or two unique cysteine residues can be constructed for specific labeling with one or two identical fluorescent probes, so-called donors (d). Fluorescence depolarization experiments on double-labeled Cys mutant monitor both reorientational motions of the d molecules, as well as the rate of intramolecular energy migration. In this report a model that accounts for these contributions to the fluorescence anisotropy is presented and experimentally tested. Mutants of a protease inhibitor, plasminogen activator inhibitor type-1 (PAI-1), containing one or two cysteine residues, were labeled with sulfhydryl specific derivatives of 4,4-difluoro-4-borata-3a-azonia-4a-aza-s-indacence (BODIPY). From the rate of energy migration, the intramolecular distance between the d groups was calculated by using the Forster mechanism and by accounting for the influence of local anisotropic orientation of the d molecules. The calculated intramolecular distances were compared with those obtained from the crystal structure of PAI-1 in its latent form. To test the stability of parameters extracted from experiments, synthetic data were generated and reanalyzed.

  • 21.
    Lidman, Martin
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Dingeldein, Artur
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Pokorná, Šárka
    Šachl, Radek
    Sparrman, Tobias
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Hof, Martin
    Gröbner, Gerhard
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    The Role of Lipids in Regulation of Programmed Cell Death2016Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 110, nr 3, s. 473A-473AArtikkel i tidsskrift (Fagfellevurdert)
  • 22. Lubart, Quentin
    et al.
    Levin, Sune
    Block, Stephan
    Joemetsa, Silver
    Kesarimangalam, Sriram
    Hook, Fredrik
    Bally, Marta
    Umeå universitet, Medicinska fakulteten, Institutionen för klinisk mikrobiologi.
    Westerlund, Fredrik
    Esbjorner, Elin
    A Nanofluidic Device for Multiplexed Analysis of Single Exosomes2020Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 118, nr 3, s. 348A-349AArtikkel i tidsskrift (Annet vitenskapelig)
  • 23.
    Mahato, Dhani R.
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Andersson, Magnus
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    State-Dependent and Mutation-Induced Differences in Protein-Lipid Interactions in the Na,K Atpase2020Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 118, nr 3, s. 497A-497AArtikkel i tidsskrift (Annet vitenskapelig)
  • 24.
    Mikaelsson, Therese
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Ådén, Jörgen
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Johansson, Lennart B-Å
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Wittung-Stafshede, Pernilla
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Direct Observation of Protein Unfolded State Compaction in the Presence of Macromolecular Crowding2013Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 104, nr 3, s. 694-704Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Proteins fold and function in cellular environments that are crowded with other macromolecules. As a consequence of excluded volume effects, compact folded states of proteins should be indirectly stabilized due to destabilization of extended unfolded conformations. Here, we assess the role of excluded volume in terms of protein stability, structural dimensions and folding dynamics using a sugar-based crowding agent, dextran 20, and the small ribosomal protein S16 as a model system. To specifically address dimensions, we labeled the protein with BODIPY at two positions and measured Trp-BODIPY distances under different conditions. As expected, we found that dextran 20 (200 mg/ml) stabilized the variants against urea-induced unfolding. At conditions where the protein is unfolded, Förster resonance energy transfer measurements reveal that in the presence of dextran, the unfolded ensemble is more compact and there is residual structure left as probed by far-ultraviolet circular dichroism. In the presence of a crowding agent, folding rates are faster in the two-state regime, and at low denaturant concentrations, a kinetic intermediate is favored. Our study provides direct evidence for protein unfolded-state compaction in the presence of macromolecular crowding along with its energetic and kinetic consequences.

  • 25.
    Mikaelsson, Therese
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Ådén, Jörgen
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Wittung-Stafshede, Pernilla
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Johansson, Lennart B-Å
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Macromolecular crowding effects on two homologs of ribosomal protein S16: protein-dependent structural changes and local interactions2014Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 107, nr 2, s. 401-410Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Proteins function in cellular environments that are crowded with biomolecules, and in this reduced available space, their biophysical properties may differ from those observed in dilute solutions in vitro. Here, we investigated the effects of a synthetic macromolecular crowding agent, dextran 20, on the folded states of hyperthermophilic (S16T(herme)) and mesophilic (S161homologs of the ribosomal protein S16. As expected for an excluded-volume effect, the resistance of the mesophilic Meso, protein to heat-induced unfolding increased in the presence of dextran 20, and chemical denaturation experiments at different fixed temperatures showed the macromolecular crowding effect to be temperature-independent. Forster resonance energy transfer experiments show that intramolecular distances between an intrinsic Trp residue and BODIPY-labeled S16 Meso depend on the level of the crowding agent. The BODIPY group was attached at three specific positions in S16me, allowing measurements of three intraprotein distances. All S16meso variants exhibited a decrease in the average Trp-BODIPY distance at up to 100 mg/mL dextran 20, whereas the changes in distance became anisotropic (one distance increased, two distances decreased) at higher dextran concentrations. In contrast, the two 516-rhermo mutants did not show any changes in Trp-BODIPY distances upon increase of dextran 20 concentrations. It should be noted that the fluorescence quantum yields and lifetimes of BODIPY attached to the two S16 homologs decreased gradually in the presence of dextran 20. To investigate the origin of this decrease, we studied the BODIPY quantum yield in three protein variants in the presence of a tyrosine-labeled dextran. The experiments revealed distinct tyrosine quenching behaviors of BODIPY in the three variants, suggesting a dynamic local interaction between dextran and one particular S16 variant.

  • 26.
    Mikhalyov, I
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Olofsson, Anders
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Gröbner, Gerhard
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Johansson, Lennart B-Å
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Designed fluorescent probes reveal interactions between Amyloid-β(1-40) Peptides and GM1 Gangliosides in Micelles and Lipid Vesicles2010Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 99, nr 5, s. 1510-1519Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    A hallmark of the common Alzheimer's disease (AD) is the pathological conversion of its amphiphatic amyloid-beta (Abeta) peptide into neurotoxic aggregates. In AD patients, these aggregates are often found to be tightly associated with neuronal G(M1) ganglioside lipids, suggesting an involvement of G(M1) not only in aggregate formation but also in neurotoxic events. Significant interactions were found between micelles made of newly synthesized fluorescent G(M1) gangliosides labeled in the polar headgroup or the hydrophobic chain and Abeta(1-40) peptide labeled with a BODIPY-FL-C1 fluorophore at positions 12 and 26, respectively. From an analysis of energy transfer between the different fluorescence labels and their location in the molecules, we were able to place the Abeta peptide inside G(M1) micelles, close to the hydrophobic-hydrophilic interface. Large unilamellar vesicles composed of a raftlike G(M1)/bSM/cholesterol lipid composition doped with labeled G(M1) at various positions also interact with labeled Abeta peptide tagged to amino acids 2 or 26. A faster energy transfer was observed from the Abeta peptide to bilayers doped with 581/591-BODIPY-C(11)-G(M1) in the nonpolar part of the lipid compared with 581/591-BODIPY-C(5)-G(M1) residing in the polar headgroup. These data are compatible with a clustering process of G(M1) molecules, an effect that not only increases the Abeta peptide affinity, but also causes a pronounced Abeta peptide penetration deeper into the lipid membrane; all these factors are potentially involved in Abeta peptide aggregate formation due to an altered ganglioside metabolism found in AD patients.

  • 27. Miller, Corwin
    et al.
    Davlieva, Milya
    Wilson, Corey
    White, Kristopher I
    Couñago, Rafael
    Wu, Gang
    Myers, Jeffrey C
    Wittung-Stafshede, Pernilla
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Shamoo, Yousif
    Experimental evolution of adenylate kinase reveals contrasting strategies toward protein thermostability2010Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 99, nr 3, s. 887-896Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Success in evolution depends critically upon the ability of organisms to adapt, a property that is also true for the proteins that contribute to the fitness of an organism. Successful protein evolution is enhanced by mutational pathways that generate a wide range of physicochemical mechanisms to adaptation. In an earlier study, we used a weak-link method to favor changes to an essential but maladapted protein, adenylate kinase (AK), within a microbial population. Six AK mutants (a single mutant followed by five double mutants) had success within the population, revealing a diverse range of adaptive strategies that included changes in nonpolar packing, protein folding dynamics, and formation of new hydrogen bonds and electrostatic networks. The first mutation, AK(BSUB) Q199R, was essential in defining the structural context that facilitated subsequent mutations as revealed by a considerable mutational epistasis and, in one case, a very strong dependence upon the order of mutations. Namely, whereas the single mutation AK(BSUB) G213E decreases protein stability by >25 degrees C, the same mutation in the background of AK(BSUB) Q199R increases stability by 3.4 degrees C, demonstrating that the order of mutations can play a critical role in favoring particular molecular pathways to adaptation. In turn, protein folding kinetics shows that four of the five AK(BSUB) double mutants utilize a strategy in which an increase in the folding rate accompanied by a decrease in the unfolding rate results in additional stability. However, one mutant exhibited a dramatic increase in the folding relative to a modest increase in the unfolding rate, suggesting a different adaptive strategy for thermostability. In all cases, an increase in the folding rates for the double mutants appears to be the preferred mechanism in conferring additional stability and may be an important aspect of protein evolution. The range of overlapping as well as contrasting strategies for success illustrates both the power and subtlety of adaptation at even the smallest unit of change, a single amino acid.

  • 28.
    Mortezaei, Narges
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för fysik.
    Singh, Bhupender
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för fysik.
    Bullitt, Esther
    Zakrisson, Johan
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för fysik.
    Epler, Chelsea
    Wiklund, Krister
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för fysik.
    Andersson, Magnus
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för fysik.
    Structural and biophysical comparison of UPEC and ETEC adhesion fimbriae2015Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 108, nr 2, suppl 1, s. 527A-527AArtikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Adhesion fimbriae (pili) of uropathogenic and enterotoxigenic Escherichia coli (UPEC and ETEC, respectively) facilitate adherence of the bacteria to target cells. Fimbriae are absolutely necessary for colonization and biofilm formation in the initiation of disease. The types of fimbriae expressed on the bacterial surface vary with the preferred environmental niche of the bacterial strain. For example, UPEC that express P-pili are most frequently associated pyelonephritis, an infection in the upper urinary tract, whereas bacteria that express type 1 fimbriae commonly cause cystitis through infection of the lower urinary tract. In contrast, ETEC expressing CFA/I and CS2 pili are associated with diarrheal diseases, initiating disease in the small intestines.

    Although expressed in different enviroments, these fimbriae share basic structural and biomechanical features. Structurally, they are all long (1-4 μm), thin (7-8 nm diameter) helix-like filaments that extend from the bacterial surface. Biomechanically, they share the ability to be extended into a thinner filament (2-3 nm diameter) by unwinding of the helical filament under a constant force. However, the force required to unwind is specific to each fimbrial type. In addition, the dependence of the force required to unwind a fimbria on the velocity of this unwinding, (that is, the kinetics of unwinding), is also type-specific and highly variable. These biomechanical parameters are dissimilar for UPEC and ETEC expressed fimbriae, separating them into two distinct groups. Using force spectroscopy data, helical reconstructions from electron microscopy data, and computational simulations, we show in this work how these pronounced biomechanical differences may be beneficial for bacterial survival in a given environment.

  • 29.
    Mortezaei, Narges
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för fysik.
    Singh, Bhupender
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för fysik.
    Uhlin, Bernt Eric
    Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten).
    Savarino, Stephen
    Bullitt, Esther
    Andersson, Magnus
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för fysik.
    Antibodies Change the Mechanics of Adhesion Fimbriae: a Case Study of CS20 Fimbriae Expressed by Enterotoxigenic Escherichia Coli2015Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 108, s. 602-Artikkel i tidsskrift (Annet vitenskapelig)
    Abstract [en]

    Enterotoxigenic Escherichia coli (ETEC) express a variety of fimbriae that mediate adhesion to host epithelial cells. It has been shown that the ability of a fimbriated bacterial cell to attach and stay attached to host cells does not merely depend on the adhesin expressed distal of the fimbriae but also the biomechanical properties of the fimbriae are vital for sustained adhesion. Fimbriae can significantly extend under a constant force when exposed to an external force and therefore reduce the load on the adhesin, which is believed to help bacteria to withstand external forces applied by various body defense systems. Thus, it is thought that the fimbrial shaft and adhesin have co-evolved for optimal function when bacteria attach to host cells. To investigate if antibodies, normally found in the intestines, affects the biomechanical properties of fimbriae, we exposed CS20 fimbriae expressed by ETEC to anti-fimbrial antibodies and measured these properties using optical tweezers force spectroscopy. Our data show a change in the force required to extend the fimbriae and that the elasticity is significantly reduced by the presence of antibodies. The reduced elasticity, likely due to cross-linking of fimbrial subunits, could thus be another assignment for antibodies; in addition to their mission in marking bacteria as foreign, our data indicate that antibodies physically compromise fimbrial function. To further confirm interaction of antibodies to their specific target we performed western blot analysis, transmission electron microscopy and immunofluoresence microscopy. In the presence of antibodies, we suggest that our assay and results will be a starting point for further studies aimed at inhibiting bacterial adhesion by antibodies.

  • 30.
    Mortezaei, Narges
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för fysik.
    Singh, Bhupender
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för fysik. Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Zakrisson, Johan
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för fysik.
    Bullitt, Esther
    Boston University School of Medicine, Boston, Massachusetts, USA.
    Andersson, Magnus
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för fysik. Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Biomechanical and Structural features of CS2 fimbriae of Enterotoxigenic Escherichia coli 2015Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 109, nr 1, s. 49-56Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Enterotoxigenic Escherichia coli (ETEC) are a major cause of diarrhea worldwide, and infection of children in underdeveloped countries often leads to high mortality rates. Isolated ETEC express a plethora of colonization factors (fimbriae/pili), of which CFA/I and CFA/II that are assembled via the alternate chaperone pathway (ACP), are amongst the most common. Fimbriae are filamentous structures, whose shafts are primarily composed of helically arranged single pilin-protein subunits, with a unique biomechanical capability allowing them to unwind and rewind. A sustained ETEC infection, under adverse conditions of dynamic shear forces, is primarily attributed to this biomechanical feature of ETEC fimbriae. Recent understandings about the role of fimbriae as virulence factors are pointing to an evolutionary adaptation of their structural and biomechanical features. In this work, we investigated the biophysical properties of CS2 fimbriae from the CFA/II group. Homology modelling its major structural subunit CotA reveals structural clues and these are related to the niche in which they are expressed. Using optical tweezers force spectroscopy we found that CS2 fimbriae unwind at a constant force of 10 pN and have a corner velocity of 1300 nm/s, i.e., the velocity at which the force required for unwinding rises exponentially with increased speed. The biophysical properties of CS2 fimbriae assessed in this work classify them into a low-force unwinding group of fimbriae together with the CFA/I and CS20 fimbriae expressed by ETEC strains. The three fimbriae are expressed by ETEC, colonize in similar gut environments, and exhibit similar biophysical features, but differ in their biogenesis. Our observation suggests that the environment has a strong impact on the biophysical characteristics of fimbriae expressed by ETEC.

  • 31. Nørregaard, Kamilla
    et al.
    Andersson, Magnus
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för fysik.
    Nielsen, Peter E
    Brown, Stanley
    Oddershede, Lene B
    DNA Supercoiling Enhances Cooperativity and Efficiency of an Epigenetic Switch2015Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 108, nr 2, s. 188A-188AArtikkel i tidsskrift (Annet vitenskapelig)
    Abstract [en]

    Upon infection by bacteriophage λ the λ repressor protein, CI, interacts with the λ operator DNA, thereby regulating protein expression and deciding between the lysogenic and lytic state. This λ switch is a model on the basis of which epigenetic switch regulation is understood. In order to study the interaction between naturally supercoiled DNA and the DNA associating protein CI, we invented a novel assay where supercoiled circular DNA plasmids were individually tethered by peptide nucleic acid (PNA) handles [1]. We used this tethered plasmid assay for a single molecule investigation of the dynamics of supercoiled DNA and studied both the dynamics of the molecule itself and its interactions with the regulatory CI protein. The dynamics of the construct was analyzed by tracking the tethered bead. This revealed that compared with relaxed DNA, the presence of supercoils greatly enhances juxtaposition probability [2]. When CI was added to the supercoiled assay, the protein would attach to the operator sites thereby looping DNA. Our studies reveal that the efficiency and cooperativity of the epigenetic λ switch are significantly increased in the supercoiled system compared with a linear assay, thus increasing the Hill coefficient [2,3]. In contrast to other single molecule assays, the current methodology allows for studying DNA dynamics and DNA-protein interactions of DNA in its naturally supercoiled state.

  • 32. Perálvarez-Marín, Alex
    et al.
    Mateos, Laura
    Zhang, Ce
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Singh, Shalini
    Cedazo-Mínguez, Angel
    Visa, Neus
    Morozova-Roche, Ludmilla
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Gräslund, Astrid
    Barth, Andreas
    Influence of residue 22 on the folding, aggregation profile, and toxicity of the Alzheimer's amyloid β peptide2009Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 97, nr 1, s. 277-285Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Several biophysical techniques have been used to determine differences in the aggregation profile (i.e., the secondary structure, aggregation propensity, dynamics, and morphology of amyloid structures) and the effects on cell viability of three variants of the amyloid beta peptide involved in Alzheimer's disease. We focused our study on the Glu22 residue, comparing the effects of freshly prepared samples and samples aged for at least 20 days. In the aged samples, a high propensity for aggregation and beta-sheet secondary structure appears when residue 22 is capable of establishing polar (Glu22 in wild-type) or hydrophobic (Val22 in E22V) interactions. The Arctic variant (E22G) presents a mixture of mostly disordered and alpha-helix structures (with low beta-sheet contribution). Analysis of transmission electron micrographs and atomic force microscopy images of the peptide variants after aging showed significant quantitative and qualitative differences in the morphology of the formed aggregates. The effect on human neuroblastoma cells of these Abeta(12-28) variants does not correlate with the amount of beta-sheet of the aggregates. In samples allowed to age, the native sequence was found to have an insignificant effect on cell viability, whereas the Arctic variant (E22G), the E22V variant, and the slightly-aggregating control (F19G-F20G) had more prominent effects.

  • 33. Preiner, Johannes
    et al.
    Janovjak, Harald
    Rankl, Christian
    Knaus, Helene
    Cisneros, David A.
    BioTec, University of Technology Dresden, Dresden, Germany.
    Kedrov, Alexej
    Kienberger, Ferry
    Muller, Daniel J
    Hinterdorfer, Peter
    Free energy of membrane protein unfolding derived from single-molecule force measurements2007Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 93, nr 3, s. 930-937Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Mechanical single-molecule techniques offer exciting possibilities to investigate protein folding and stability in native environments at submolecular resolution. By applying a free-energy reconstruction procedure developed by Hummer and Szabo, which is based on a statistical theorem introduced by Jarzynski, we determined the unfolding free energy of the membrane proteins bacteriorhodopsin (BR), halorhodopsin, and the sodium-proton antiporter NhaA. The calculated energies ranged from 290.5 kcal/mol for BR to 485.5 kcal/mol for NhaA. For the remarkably stable BR, the equilibrium unfolding free energy was independent of pulling rate and temperature ranging between 18 and 42 degrees C. Our experiments also revealed heterogeneous energetic properties in individual transmembrane helices. In halorhodopsin, the stabilization of a short helical segment yielded a characteristic signature in the energy profile. In NhaA, a pronounced peak was observed at a functionally important site in the protein. Since a large variety of single- and multispan membrane proteins can be tackled in mechanical unfolding experiments, our approach provides a basis for systematically elucidating energetic properties of membrane proteins with the resolution of individual secondary-structure elements.

  • 34.
    Ravishankar, Harsha
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Goodman, Jack
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Nors Pedersen, Martin
    Wulff, Michael
    Levantino, Matteo
    Wolf-Watz, Magnus
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Andersson, Magnus
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Mapping the Adenylate Kinase Reaction by Time-Resolved X-ray Solution Scattering2020Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 118, nr 3, s. 50A-50AArtikkel i tidsskrift (Annet vitenskapelig)
  • 35.
    Ravishankar, Harsha
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Pedersen, Martin Nors
    Sitsel, Alya
    Li, Chenge
    Duelli, Annette
    Levantino, Matteo
    Wulff, Michael
    Barth, Andreas
    Olesen, Claus
    Nissen, Poul
    Andersson, Magnus
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Tracking Ca2+ ATPase Intermediates in Real-time by X-Ray Solution Scattering2020Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 118, nr 3, s. 26A-26AArtikkel i tidsskrift (Annet vitenskapelig)
  • 36.
    Rogne, Per
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Sauer-Eriksson, Elisabeth
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Sauer, Uwe H.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Hedberg, Christian
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Wolf-Watz, Magnus
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Principles of ATP and GTP Selectivity in NMP Kinases2020Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 118, nr 3, s. 193A-193AArtikkel i tidsskrift (Annet vitenskapelig)
  • 37.
    Rogne, Per
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Wolf-Watz, Magnus
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Urea-Dependent Adenylate Kinase Activation following Redistribution of Structural States2016Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 111, nr 7, s. 1385-1395Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Proteins are often functionally dependent on conformational changes that allow them to sample structural states that are sparsely populated in the absence of a substrate or binding partner. The distribution of such structural microstates is governed by their relative stability, and the kinetics of their interconversion is governed by the magnitude of associated activation barriers. Here, we have explored the interplay among structure, stability, and function of a selected enzyme, adenylate kinase (Adk), by monitoring changes in its enzymatic activity in response to additions of urea. For this purpose we used a 31P NMR assay that was found useful for heterogeneous sample compositions such as presence of urea. It was found that Adk is activated at low urea concentrations whereas higher urea concentrations unfolds and thereby deactivates the enzyme. From a quantitative analysis of chemical shifts, it was found that urea redistributes preexisting structural microstates, stabilizing a substrate-bound open state at the expense of a substrate-bound closed state. Adk is rate-limited by slow opening of substrate binding domains and the urea-dependent redistribution of structural states is consistent with a model where the increased activity results from an increased rate-constant for domain opening. In addition, we also detected a strong correlation between the catalytic free energy and free energy of substrate (ATP) binding, which is also consistent with the catalytic model for Adk. From a general perspective, it appears that urea can be used to modulate conformational equilibria of folded proteins toward more expanded states for cases where a sizeable difference in solvent-accessible surface area exists between the states involved. This effect complements the action of osmolytes, such as trimethylamine N-oxide, that favor more compact protein states.

  • 38. Rojdestvenski, I
    et al.
    Ivanov, A G
    Cottam, M G
    Borodich, A
    Huner, N P A
    Oquist, Gunnar
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för fysiologisk botanik. Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Umeå Plant Science Centre (UPSC).
    Segregation of photosystems in thylakoid membranes as a critical phenomenon2002Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 82, nr 4, s. 1719-1730Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The distribution of the two photosystems, PSI and PSII, in grana and stroma lamellae of the chloroplast membranes is not uniform. PSII are mainly concentrated in grana and PSI in stroma thylakoids. The dynamics and factors controlling the spatial segregation of PSI and PSII are generally not well understood, and here we address the segregation of photosystems in thylakoid membranes by means of a molecular dynamics method. The lateral segregation of photosystems was studied assuming a model comprising a two-dimensional (in-plane), two-component, many-body system with periodic boundary conditions and competing interactions between the photosystems in the thylakoid membrane. PSI and PSII are represented by particles with different values of negative charge. The pair interactions between particles include a screened Coulomb repulsive part and an exponentially decaying attractive part. The modeling results suggest a complicated phase behavior of the system, including quasi-crystalline phase of randomly distributed complexes of PSII and PSI at low ionic screening, well defined clustered state of segregated complexes at high screening, and in addition, an intermediate agglomerate phase where the photosystems tend to aggregate together without segregation. The calculations demonstrated that the ordering of photosystems within the membrane was the result of interplay between electrostatic and lipid-mediated interactions. At some values of the model parameters the segregation can be represented visually as well as by analyzing the correlation functions of the configuration.

  • 39.
    Sachl, Radek
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Humpolickova, Jana
    J. Heyrovský Institute of Physical Chemistry, v.v.i., Academy of Sciences of the Czech Republic, Prague, Czech Republic.
    Stefl, Martin
    J. Heyrovský Institute of Physical Chemistry, v.v.i., Academy of Sciences of the Czech Republic, Prague, Czech Republic.
    Johansson, Lennart B-Å
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Hof, Martin
    J. Heyrovsky´ Institute of Physical Chemistry, v.v.i., Academy of Sciences of the Czech Republic, Prague, Czech Republic.
    Limitations of electronic energy transfer in the determination of lipid nanodomain sizes2011Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 101, nr 11, s. L60-L62Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Even though superresolution microscopy indicates that size of plasma membrane rafts is <20 nm, those structures have never been observed. Forster resonance energy transfer (FRET) is therefore still the most powerful optical method for characterization of such domains. In this letter we investigate relation between nanodomain affinity of a donor-acceptor (D/A) pair and the detectable nanodomain size/area. We show that probes with high affinity to the liquid-ordered (L(o)) phase are required for detecting domain sizes of a few nanometers, and/or domains that occupy a few percent of the bilayer area. A combination of donors and acceptors that prefer different phases is the more favorable approach. For instance, a D/A pair with the distribution constant of donors K(D) = 5 and acceptors K(A) = 0.01 can resolve a broad spectrum of nanodomain sizes. On the other hand, currently available donors and acceptors that prefer the same phase, either the liquid-disordered (L(d)) or L(o) phase, are not so convenient for determining domain sizes <20 nm. Here the detection limits of FRET experiments employing several commonly used D/A pairs have been investigated.

  • 40.
    Stefl, Martin
    et al.
    Acad Sci Czech Republic, J Heyrovsky Inst Phys Chem, Dept Biophys Chem, Prague, Czech Republic .
    Sachl, Radek
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Humpolickova, Jana
    Acad Sci Czech Republic, J Heyrovsky Inst Phys Chem, Dept Biophys Chem, Prague, Czech Republic .
    Cebecauer, Marek
    Acad Sci Czech Republic, J Heyrovsky Inst Phys Chem, Dept Biophys Chem, Prague, Czech Republic .
    Machan, Radek
    Acad Sci Czech Republic, J Heyrovsky Inst Phys Chem, Dept Biophys Chem, Prague, Czech Republic .
    Kolarova, Marie
    Acad Sci Czech Republic, J Heyrovsky Inst Phys Chem, Dept Biophys Chem, Prague, Czech Republic .
    Johansson, Lennart B-Å
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Hof, Martin
    Acad Sci Czech Republic, J Heyrovsky Inst Phys Chem, Dept Biophys Chem, Prague, Czech Republic .
    Dynamics and size of cross-linking-induced lipid Nanodomains in model Membranes2012Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 102, nr 9, s. 2104-2113Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Changes of membrane organization upon cross-linking of its components trigger cell signaling response to various exogenous factors. Cross-linking of raft gangliosides GM1 with cholera toxin ( CTxB) was shown to cause microscopic phase separation in model membranes, and the CTxB-GM1 complexes forming a minimal lipid raft unit are the subject of ongoing cell membrane research. Yet, those subdiffraction sized rafts have never been described in terms of size and dynamics. By means of two-color z-scan fluorescence correlation spectroscopy, we show that the nanosized domains are formed in model membranes at lower sphingomyelin (Sph) content than needed for the large-scale phase separation and that the CTxB-GM1 complexes are confined in the domains poorly stabilized with Sph. Forster resonance energy transfer together with Monte Carlo modeling of the donor decay response reveal the domain radius of similar to 8 nm, which increases at higher Sph content. We observed two types of domains behaving differently, which suggests a dual role of the cross-linker: first, local transient condensation of the GM1 molecules compensating for a lack of Sph and second, coalescence of existing nanodomains ending in large-scale phase separation.

  • 41.
    Vestergren, Johan E.
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Vincent, Andrea G.
    Persson, Per
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Jansson, Mats
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för ekologi, miljö och geovetenskap.
    Ilstedt, Ulrik
    Giesler, Reiner
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för ekologi, miljö och geovetenskap.
    Schleucher, Jurgen
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Gröbner, Gerhard
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Novel Approaches for Identifying Phosphorus Species in Terrestrial and Aquatic Ecosystems with P-31 NMR2013Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 104, nr 2, s. 501A-502AArtikkel i tidsskrift (Annet vitenskapelig)
  • 42.
    Wallgren, Martin
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Pham, Quoc Dat
    Lidman, Martin
    Kinnunen, Paavo K. J.
    Hof, Martin
    Gröbner, Gerhard
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Impact of Oxidized Phospholipids on Membrane Organization2013Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 104, nr 2, s. 249A-249AArtikkel i tidsskrift (Annet vitenskapelig)
  • 43.
    Weise, Christoph F
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Login, Frédéric H
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Ho, Oanh
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Gröbner, Gerhard
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Wolf-Watz, Hans
    Umeå universitet, Medicinska fakulteten, Institutionen för molekylärbiologi (Medicinska fakulteten). Umeå universitet, Medicinska fakulteten, Molekylär Infektionsmedicin, Sverige (MIMS). Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Wolf-Watz, Magnus
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Negatively charged lipid membranes promote a disorder-order transition in the Yersinia YscU protein2014Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 107, nr 8, s. 1950-1961Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    The inner membrane of Gram-negative bacteria is negatively charged, rendering positively charged cytoplasmic proteins in close proximity likely candidates for protein-membrane interactions. YscU is a Yersinia pseudotuberculosis type III secretion system protein crucial for bacterial pathogenesis. The protein contains a highly conserved positively charged linker sequence that separates membrane-spanning and cytoplasmic (YscUC) domains. Although disordered in solution, inspection of the primary sequence of the linker reveals that positively charged residues are separated with a typical helical periodicity. Here, we demonstrate that the linker sequence of YscU undergoes a largely electrostatically driven coil-to-helix transition upon binding to negatively charged membrane interfaces. Using membrane-mimicking sodium dodecyl sulfate micelles, an NMR derived structural model reveals the induction of three helical segments in the linker. The overall linker placement in sodium dodecyl sulfate micelles was identified by NMR experiments including paramagnetic relaxation enhancements. Partitioning of individual residues agrees with their hydrophobicity and supports an interfacial positioning of the helices. Replacement of positively charged linker residues with alanine resulted in YscUC variants displaying attenuated membrane-binding affinities, suggesting that the membrane interaction depends on positive charges within the linker. In vivo experiments with bacteria expressing these YscU replacements resulted in phenotypes displaying significantly reduced effector protein secretion levels. Taken together, our data identify a previously unknown membrane-interacting surface of YscUC that, when perturbed by mutations, disrupts the function of the pathogenic machinery in Yersinia.

  • 44. Williamson, Philip T. F.
    et al.
    Horrocks, Jack
    Maheswaran, Luckshi
    Concistre, Maria
    Morozova-Roche, Ludmilla
    Umeå universitet, Medicinska fakulteten, Institutionen för medicinsk kemi och biofysik.
    Unravelling the Role of S100A9 in the Development of Neurodegenerative Disease2019Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 116, nr 3, s. 338A-338AArtikkel i tidsskrift (Annet vitenskapelig)
  • 45.
    Wittung-Stafshede, Pernilla
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    In Vitro effects of Macromolecular Crowding on Protein Stability, Structure and Folding2013Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 104, nr 2, s. 576A-576AArtikkel i tidsskrift (Annet vitenskapelig)
  • 46.
    Wittung-Stafshede, Pernilla
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Protein folding inside the cell2011Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 101, nr 2, s. 265-266Artikkel i tidsskrift (Fagfellevurdert)
  • 47.
    Wittung-Stafshede, Pernilla
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Tuning of Alpha-Synuclein Aggregation by Small Molecules and Bacterial Proteins2015Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 108, nr 2, s. 522A-522AArtikkel i tidsskrift (Annet vitenskapelig)
    Abstract [en]

    Parkinson's disease affects a growing number of the population and involves motor complications due to the death of dopamine neurons. Cytosolic inclusions containing amyloid fibrils of α-synuclein are a hallmark of the disease and it is believed that the aggregation process (going from monomers to amyloid fibers) of alpha-synculein somehow causes neurodegeneration. The synuclein-rich inclusions share structural characteristics with amyloid fibers found in many other neurodegenerative disorders. In addition, many organisms employ amyloid structures for mechanical or biological functions; for example, amyloid fibers are the major component of microbial biofilms. Mature amyloid fibers of alpha-synculein may not be the source of cytotoxicity; instead, transient oligomeric structures may be most dangerous to the neuronal cells. To investigate molecular pathways leading to alpha-synculein amyloid fibers, and thereby get hints for how to combat Parkinson's disease in vivo, we have taken a unique approach that involves purified proteins, biophysical experiments in vitro, and small-molecule tools. We have found that strategic ring-fused 2-pyridone compounds (mimics of small peptides), can tune alpha-synuclein aggregation such that either inhibitory or templating oligomers accumulate. Moreover, a fine balance between templation and inhibition processes is evident since one particular 2-pyridone inhibits bacterial amyloid formation but promotes alpha-synuclein amyloid fibers. In analogy with the small molecule tools, we found that bacterial proteins can cross-react with alpha-synculein and inhibit as well as promote amyloid fiber formation at sub-stoichiometric levels. Direct interactions of alpha-synculein with bacterial proteins and/or natural metabolites may play a role in controlling Parkinson's disease in humans.

  • 48.
    Wolf-Watz, Magnus
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Kovermann, Michael
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Dynamics of a Naturally Hidden State Restricts Adenylate Kinase Activity2015Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 108, nr 2, s. 30A-30AArtikkel i tidsskrift (Annet vitenskapelig)
  • 49.
    Wolf-Watz, Magnus
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Rogne, Per
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Sauer-Eriksson, A. Elisabeth
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Sauer, Uwe H.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Hedberg, Christian
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Kemiska institutionen.
    Positive and Negative Substrate Interference Supported by Coinciding Enzyme Residues2019Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 116, nr 3, s. 485A-485AArtikkel i tidsskrift (Annet vitenskapelig)
  • 50.
    Zakrisson, Johan
    et al.
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för fysik. Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Wiklund, Krister
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för fysik. Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Axner, Ove
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för fysik. Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    Andersson, Magnus
    Umeå universitet, Teknisk-naturvetenskapliga fakulteten, Institutionen för fysik. Umeå universitet, Medicinska fakulteten, Umeå Centre for Microbial Research (UCMR).
    The shaft of the type 1 fimbriae regulates an externalforce to match the FimH catch bond2013Inngår i: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 104, nr 10, s. 2137-2148Artikkel i tidsskrift (Fagfellevurdert)
    Abstract [en]

    Type 1 fimbriae mediate adhesion of uropathogenic Escherichia coli to host cells. It has been hypothesized that due to their ability to uncoil under exposure to force, fimbriae can reduce fluid shear stress on the adhesin-receptor interaction by which the bacterium adheres to the surface. In this work, we develop a model that describes how the force on the adhesin-receptor interaction of a type 1 fimbria varies as a bacterium is affected by a time-dependent fluid flow mimicking in vivo conditions. The model combines in vivo hydrodynamic conditions with previously assessed biomechanical properties of the fimbriae. Numerical methods are used to solve for the motion and adhesion force under the presence of time-dependent fluid profiles. It is found that a bacterium tethered with a type 1 pilus will experience significantly reduced shear stress for moderate to high flow velocities and that the maximum stress the adhesin will experience is limited to ∼120 pN, which is sufficient to activate the conformational change of the FimH adhesin into its stronger state but also lower than the force required for breaking it under rapid loading. Our model thus supports the assumption that the type 1 fimbria shaft and the FimH adhesin-receptor interaction are optimized to each other, and that they give piliated bacteria significant advantages in rapidly changing fluidic environments.

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