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Jonna, Venkateswara Rao
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Publications (10 of 14) Show all publications
Gupta, A. A., Reinartz, I., Karunanithy, G., Spilotros, A., Jonna, V. R., Hofer, A., . . . Wolf-Watz, M. (2018). Formation of a Secretion-Competent Protein Complex by a Dynamic Wrap-around Binding Mechanism. Journal of Molecular Biology, 430(18, Part B), 3157-3169
Open this publication in new window or tab >>Formation of a Secretion-Competent Protein Complex by a Dynamic Wrap-around Binding Mechanism
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2018 (English)In: Journal of Molecular Biology, ISSN 0022-2836, E-ISSN 1089-8638, Vol. 430, no 18, Part B, p. 3157-3169Article in journal (Refereed) Published
Abstract [en]

Bacterial virulence is typically initiated by translocation of effector or toxic proteins across host cell membranes. A class of gram-negative pathogenic bacteria including Yersinia pseudotuberculosis and Yersinia pestis accomplishes this objective with a protein assembly called the type III secretion system. Yersinia effector proteins (Yop) are presented to the translocation apparatus through formation of specific complexes with their cognate chaperones (Syc). In the complexes where the structure is available, the Yops are extended and wrap around their cognate chaperone. This structural architecture enables secretion of the Yop from the bacterium in early stages of translocation. It has been shown previously that the chaperone-binding domain of YopE is disordered in its isolation but becomes substantially more ordered in its wrap-around complex with its chaperone SycE. Here, by means of NMR spectroscopy, small-angle X-ray scattering and molecular modeling, we demonstrate that while the free chaperone-binding domain of YopH (YopHCBD) adopts a fully ordered and globular fold, it populates an elongated, wrap-around conformation when it engages in a specific complex with its chaperone SycH2. Hence, in contrast to YopE that is unstructured in its free state, YopH transits from a globular free state to an elongated chaperone-bound state. We demonstrate that a sparsely populated YopHCBD state has an elevated affinity for SycH2 and represents an intermediate in the formation of the protein complex. Our results suggest that Yersinia has evolved a binding mechanism where SycH2 passively stimulates an elongated YopH conformation that is presented to the type III secretion system in a secretion-competent conformation.

Place, publisher, year, edition, pages
Elsevier, 2018
Keywords
NMR spectroscopy, protein complex, binding mechanism
National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:umu:diva-151495 (URN)10.1016/j.jmb.2018.07.014 (DOI)000444668100010 ()
Funder
Swedish Research Council, 2013-5954Knut and Alice Wallenberg FoundationCarl Tryggers foundation , CTS 15:210The Kempe Foundations
Available from: 2018-09-05 Created: 2018-09-05 Last updated: 2018-10-04Bibliographically approved
Rozman Grinberg, I., Lundin, D., Hasan, M., Crona, M., Rao Jonna, V., Loderer, C., . . . Sjöberg, B.-M. (2018). Novel ATP-cone-driven allosteric regulation of ribonucleotide reductase via the radical-generating subunit. eLIFE, 7, Article ID e31529.
Open this publication in new window or tab >>Novel ATP-cone-driven allosteric regulation of ribonucleotide reductase via the radical-generating subunit
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2018 (English)In: eLIFE, E-ISSN 2050-084X, Vol. 7, article id e31529Article in journal (Refereed) Published
Abstract [en]

Ribonucleotide reductases (RNRs) are key enzymes in DNA metabolism, with allosteric mechanisms controlling substrate specificity and overall activity. In RNRs, the activity master-switch, the ATP-cone, has been found exclusively in the catalytic subunit. In two class I RNR subclasses whose catalytic subunit lacks the ATP-cone, we discovered ATP-cones in the radical-generating subunit. The ATP-cone in the Leeuwenhoekiella blandensis radical-generating subunit regulates activity via quaternary structure induced by binding of nucleotides. ATP induces enzymatically competent dimers, whereas dATP induces non-productive tetramers, resulting in different holoenzymes. The tetramer forms by interactions between ATP-cones, shown by a 2.45 A crystal structure. We also present evidence for an (MnMnIV)-Mn-III metal center. In summary, lack of an ATP-cone domain in the catalytic subunit was compensated by transfer of the domain to the radical-generating subunit. To our knowledge, this represents the first observation of transfer of an allosteric domain between components of the same enzyme complex.

Place, publisher, year, edition, pages
ELIFE SCIENCES PUBLICATIONS LTD, 2018
National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:umu:diva-144937 (URN)10.7554/eLife.31529 (DOI)000423786200001 ()
Available from: 2018-02-23 Created: 2018-02-23 Last updated: 2018-06-09Bibliographically approved
Grinberg, I., McGann, M., Lundin, D., Crona, M., Hasan, M., Jonna, V. R., . . . Sjöberg, B.-M. (2018). Novel ATP-Cone-Driven Allosteric Regulation of Ribonucleotide Reductase Via the Radical-Generating Subunit. Paper presented at 32nd Annual Symposium of the Protein-Society, JUL 09-12, 2018, Boston, MA. Protein Science, 27, 87-88
Open this publication in new window or tab >>Novel ATP-Cone-Driven Allosteric Regulation of Ribonucleotide Reductase Via the Radical-Generating Subunit
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2018 (English)In: Protein Science, ISSN 0961-8368, E-ISSN 1469-896X, Vol. 27, p. 87-88Article in journal, Meeting abstract (Other academic) Published
Place, publisher, year, edition, pages
Wiley-Blackwell, 2018
National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:umu:diva-154065 (URN)000450682700147 ()
Conference
32nd Annual Symposium of the Protein-Society, JUL 09-12, 2018, Boston, MA
Available from: 2018-12-19 Created: 2018-12-19 Last updated: 2018-12-19Bibliographically approved
Schmitt, A., Jiang, K., Camacho, M. I., Jonna, V. R., Hofer, A., Westerlund, F., . . . Berntsson, R.-A. P. (2018). PrgB promotes aggregation, biofilm formation, and conjugation through DNA binding and compaction. Molecular Microbiology, 109(3), 291-305
Open this publication in new window or tab >>PrgB promotes aggregation, biofilm formation, and conjugation through DNA binding and compaction
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2018 (English)In: Molecular Microbiology, ISSN 0950-382X, E-ISSN 1365-2958, Vol. 109, no 3, p. 291-305Article in journal (Refereed) Published
Abstract [en]

Gram-positive bacteria deploy type IV secretion systems (T4SSs) to facilitate horizontal gene transfer. The T4SSs of Gram-positive bacteria rely on surface adhesins as opposed to conjugative pili to facilitate mating. Enterococcus faecalis PrgB is a surface adhesin that promotes mating pair formation and robust biofilm development in an extracellular DNA (eDNA) dependent manner. Here, we report the structure of the adhesin domain of PrgB. The adhesin domain binds and compacts DNA in vitro. In vivo PrgB deleted of its adhesin domain does not support cellular aggregation, biofilm development and conjugative DNA transfer. PrgB also binds lipoteichoic acid (LTA), which competes with DNA binding. We propose that PrgB binding and compaction of eDNA facilitates cell aggregation and plays an important role in establishment of early biofilms in mono- or polyspecies settings. Within these biofilms, PrgB mediates formation and stabilization of direct cell-cell contacts through alternative binding of cell-bound LTA, which in turn promotes establishment of productive mating junctions and efficient intra- or inter-species T4SS-mediated gene transfer.

Place, publisher, year, edition, pages
John Wiley & Sons, 2018
National Category
Microbiology in the medical area
Identifiers
urn:nbn:se:umu:diva-152231 (URN)10.1111/mmi.13980 (DOI)000443810300005 ()29723434 (PubMedID)
Available from: 2018-10-25 Created: 2018-10-25 Last updated: 2018-10-25Bibliographically approved
Loderer, C., Jonna, V. R., Crona, M., Grinberg, I. R., Sahlin, M., Hofer, A., . . . Sjoeberg, B.-M. (2017). A unique cysteine-rich zinc finger domain present in a majority of class II ribonucleotide reductases mediates catalytic turnover. Journal of Biological Chemistry, 292(46), 19044-19054
Open this publication in new window or tab >>A unique cysteine-rich zinc finger domain present in a majority of class II ribonucleotide reductases mediates catalytic turnover
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2017 (English)In: Journal of Biological Chemistry, ISSN 0021-9258, E-ISSN 1083-351X, Vol. 292, no 46, p. 19044-19054Article in journal (Refereed) Published
Abstract [en]

Ribonucleotide reductases (RNRs) catalyze the reduction of ribonucleotides to the corresponding deoxyribonucleotides, used in DNA synthesis and repair. Two different mechanisms help deliver the required electrons to the RNR active site. Formate can be used as reductant directly in the active site, or glutaredoxins or thioredoxins reduce a C-terminal cysteine pair, which then delivers the electrons to the active site. Here, we characterized a novel cysteine-rich C-terminal domain (CRD), which is present in most class II RNRs found in microbes. The NrdJd-type RNR from the bacterium Stackebrandtia nassauensis was used as a model enzyme. We show that the CRD is involved in both higher oligomeric state formation and electron transfer to the active site. The CRD-dependent formation of high oligomers, such as tetramers and hexamers, was induced by addition of dATP or dGTP, but not of dTTP or dCTP. The electron transfer was mediated by an array of six cysteine residues at the very C-terminal end, which also coordinated a zinc atom. The electron transfer can also occur between subunits, depending on the enzyme's oligomeric state. An investigation of the native reductant of the system revealed no interaction with glutaredoxins or thioredoxins, indicating that this class II RNR uses a different electron source. Our results indicate that the CRD has a crucial role in catalytic turnover and a potentially new terminal reduction mechanism and suggest that the CRD is important for the activities of many class II RNRs.

Place, publisher, year, edition, pages
AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC, 2017
Keywords
metal ion-protein interaction, oligomerization, oxidation-reduction (redox), phylogenetics, ribonucleotide reductase, thioredoxin
National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:umu:diva-142970 (URN)10.1074/jbc.M117.806331 (DOI)000415848000027 ()28972190 (PubMedID)
Available from: 2017-12-14 Created: 2017-12-14 Last updated: 2018-06-09Bibliographically approved
Jonna, V. R. (2017). Class I Ribonucleotide Reductases: overall activity regulation, oligomerization, and drug targeting. (Doctoral dissertation). Umeå: Umeå universitet
Open this publication in new window or tab >>Class I Ribonucleotide Reductases: overall activity regulation, oligomerization, and drug targeting
2017 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Ribonucleotide reductase (RNR) is a key enzyme in the de novo biosynthesis and homeostatic maintenance of all four DNA building blocks by being able to make deoxyribonucleotides from the corresponding ribonucleotides. It is important for the cell to control the production of a balanced supply of the dNTPs to minimize misincorporations in DNA. Because RNR is the rate-limiting enzyme in DNA synthesis, it is an important target for antimicrobial and antiproliferative molecules. The enzyme RNR has one of the most sophisticated allosteric regulations known in Nature with four allosteric effectors (ATP, dATP, dGTP, and dTTP) and two allosteric sites. One of the sites (s-site) controls the substrate specificity of the enzyme, whereas the other one (a-site) regulates the overall activity.  The a-site binds either dATP, which inhibits the enzyme or ATP that activates the enzyme. In eukaryotes, ATP activation is directly through the a-site and in E. coli it is a cross-talk effect between the a and s-sites. It is important to study and get more knowledge about the overall activity regulation of RNR, both because it has an important physiological function, but also because it may provide important clues to the design of antibacterial and antiproliferative drugs, which can target RNR.

Previous studies of class I RNRs, the class found in nearly all eukaryotes and many prokaryotes have revealed that the overall activity regulation is dependent on the formation of oligomeric complexes. The class I RNR consists of two subunits, a large α subunit, and a small β subunit. The oligomeric complexes vary between different species with the mammalian and yeast enzymes cycle between structurally different active and inactive α6β2 complexes, and the E. coli enzyme cycles between active α2β2 and inactive α4β4 complexes. Because RNR equilibrates between many different oligomeric forms that are not resolved by most conventional methods, we have used a technique termed gas-phase electrophoretic macromolecule analysis (GEMMA). In the present studies, our focus is on characterizing both prokaryotic and mammalian class I RNRs. In one of our projects, we have studied the class I RNR from Pseudomonas aeruginosa and found that it represents a novel mechanism of overall activity allosteric regulation, which is different from the two known overall activity allosteric regulation found in E. coli and eukaryotic RNRs, respectively.  The structural differences between the bacterial and the eukaryote class I RNRs are interesting from a drug developmental viewpoint because they open up the possibility of finding inhibitors that selectively target the pathogens. The biochemical data that we have published in the above project was later supported by crystal structure and solution X-ray scattering data that we published together with Derek T. Logan`s research group.

We have also studied the effect of a novel antiproliferative molecule, NSC73735, on the oligomerization of the human RNR large subunit. This collaborative research results showed that the molecule NSC73735 is the first reported non-nucleoside molecule which alters the oligomerization to inhibit human RNR and the molecule disrupts the cell cycle distribution in human leukemia cells.

Place, publisher, year, edition, pages
Umeå: Umeå universitet, 2017. p. 51+8
Series
Umeå University medical dissertations, ISSN 0346-6612 ; 1894
Keywords
Ribonucleotide reductase, GEMMA, Allosteric regulation
National Category
Biochemistry and Molecular Biology
Research subject
Medical Biochemistry
Identifiers
urn:nbn:se:umu:diva-133817 (URN)978-91-7601-703-6 (ISBN)
Public defence
2017-05-12, KB.E3.01 Lilla Hörsalen, KBC huset, Umeå, 09:00 (English)
Opponent
Supervisors
Available from: 2017-04-21 Created: 2017-04-18 Last updated: 2018-06-09Bibliographically approved
Gupta, A., Reinartz, I., Spilotros, A., Jonna, V. R., Hofer, A., Svergun, D. I., . . . Wolf-Watz, M. (2017). Global Disordering in Stereo-Specific Protein Association. Paper presented at 61st Annual Meeting of the Biophysical-Society, FEB 11-15, 2017, New Orleans, LA. Biophysical Journal, 112(3), 33A-33A
Open this publication in new window or tab >>Global Disordering in Stereo-Specific Protein Association
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2017 (English)In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 112, no 3, p. 33A-33AArticle in journal, Meeting abstract (Refereed) Published
Place, publisher, year, edition, pages
CELL PRESS, 2017
National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Identifiers
urn:nbn:se:umu:diva-137015 (URN)000402328000166 ()
Conference
61st Annual Meeting of the Biophysical-Society, FEB 11-15, 2017, New Orleans, LA
Available from: 2017-06-29 Created: 2017-06-29 Last updated: 2018-06-09Bibliographically approved
Crona, M., Codo, P., Jonna, V. R., Hofer, A., Fernandes, A. P. & Tholander, F. (2016). A ribonucleotide reductase inhibitor with deoxyribonucleoside-reversible cytotoxicity. Molecular Oncology, 10(9), 1375-1386
Open this publication in new window or tab >>A ribonucleotide reductase inhibitor with deoxyribonucleoside-reversible cytotoxicity
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2016 (English)In: Molecular Oncology, ISSN 1574-7891, E-ISSN 1878-0261, Vol. 10, no 9, p. 1375-1386Article in journal (Refereed) Published
Abstract [en]

Ribonucleotide Reductase (RNR) is the sole enzyme that catalyzes the reduction of ribonucleotides into deoxyribonucleotides. Even though RNR is a recognized target for antiproliferative molecules, and the main target of the approved drug hydroxyurea, few new leads targeted to this enzyme have been developed. We have evaluated a recently identified set of RNR inhibitors with respect to inhibition of the human enzyme and cellular toxicity. One compound, NSC73735, is particularly interesting; it is specific for leukemia cells and is the first identified compound that hinders oligomerization of the mammalian large RNR subunit. Similar to hydroxyurea, it caused a disruption of the cell cycle distribution of cultured HL-60 cells. In contrast to hydroxyurea, the disruption was reversible, indicating higher specificity. NSC73735 thus defines a potential lead candidate for RNR-targeted anticancer drugs, as well as a chemical probe with better selectivity for RNR inhibition than hydroxyurea. 

Keywords
Ribonucleotide reductase, Nucleotide metabolism, Inhibitors, Antiproliferative compounds, Cell cycle, Cytotoxicity, Oligomeric state, GEMMA
National Category
Cell and Molecular Biology Cancer and Oncology
Identifiers
urn:nbn:se:umu:diva-129926 (URN)10.1016/j.molonc.2016.07.008 (DOI)000386860800001 ()27511871 (PubMedID)
Available from: 2017-01-10 Created: 2017-01-10 Last updated: 2018-06-09Bibliographically approved
Chiruvella, K. K., Rajaei, N., Jonna, V. R., Hofer, A. & Åstrom, S. U. (2016). Biochemical Characterization of Kat1: a Domesticated hAT-Transposase that Induces DNA Hairpin Formation and MAT-Switching. Scientific Reports, 6, Article ID 21671.
Open this publication in new window or tab >>Biochemical Characterization of Kat1: a Domesticated hAT-Transposase that Induces DNA Hairpin Formation and MAT-Switching
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2016 (English)In: Scientific Reports, ISSN 2045-2322, E-ISSN 2045-2322, Vol. 6, article id 21671Article in journal (Refereed) Published
Abstract [en]

Kluyveromyces lactis hAT-transposase 1 (Kat1) generates hairpin-capped DNA double strand breaks leading to MAT-switching (MATa to MAT alpha). Using purified Kat1, we demonstrate the importance of terminal inverted repeats and subterminal repeats for its endonuclease activity. Kat1 promoted joining of the transposon end into a target DNA molecule in vitro, a biochemical feature that ties Kat1 to transposases. Gas-phase Electrophoretic Mobility Macromolecule analysis revealed that Kat1 can form hexamers when complexed with DNA. Kat1 point mutants were generated in conserved positions to explore structure-function relationships. Mutants of predicted catalytic residues abolished both DNA cleavage and strand-transfer. Interestingly, W576A predicted to be impaired for hairpin formation, was active for DNA cleavage and supported wild type levels of mating-type switching. In contrast, the conserved CXXH motif was critical for hairpin formation because Kat1 C402A/H405A completely blocked hairpinning and switching, but still generated nicks in the DNA. Mutations in the BED zinc-finger domain (C130A/C133A) resulted in an unspecific nuclease activity, presumably due to nonspecific DNA interaction. Kat1 mutants that were defective for cleavage in vitro were also defective for mating-type switching. Collectively, this study reveals Kat1 sharing extensive biochemical similarities with cut and paste transposons despite being domesticated and evolutionary diverged from active transposons.

Place, publisher, year, edition, pages
Nature Publishing Group, 2016
National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Identifiers
urn:nbn:se:umu:diva-118247 (URN)10.1038/srep21671 (DOI)000370630000001 ()
Available from: 2016-03-17 Created: 2016-03-14 Last updated: 2018-06-07Bibliographically approved
Johansson, R., Jonna, V. R., Kumar, R., Nayeri, N., Lundin, D., Sjöberg, B.-M., . . . Logan, D. T. (2016). Structural Mechanism of Allosteric Activity Regulation in a Ribonucleotide Reductase with Double ATP Cones. Structure, 24(6), 906-917
Open this publication in new window or tab >>Structural Mechanism of Allosteric Activity Regulation in a Ribonucleotide Reductase with Double ATP Cones
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2016 (English)In: Structure, ISSN 0969-2126, E-ISSN 1878-4186, Vol. 24, no 6, p. 906-917Article in journal (Refereed) Published
Abstract [en]

Ribonucleotide reductases (RNRs) reduce ribonucleotides to deoxyribonucleotides. Their overall activity is stimulated by ATP and downregulated by dATP via a genetically mobile ATP cone domain mediating the formation of oligomeric complexes with varying quaternary structures. The crystal structure and solution X-ray scattering data of a novel dATP-induced homotetramer of the Pseudomonas aeruginosa class I RNR reveal the structural bases for its unique properties, namely one ATP cone that binds two dATP molecules and a second one that is non-functional, binding no nucleotides. Mutations in the observed tetramer interface ablate oligomerization and dATP-induced inhibition but not the ability to bind dATP. Sequence analysis shows that the novel type of ATP cone may be widespread in RNRs. The present study supports a scenario in which diverse mechanisms for allosteric activity regulation are gained and lost through acquisition and evolutionary erosion of different types of ATP cone.

National Category
Cell and Molecular Biology
Identifiers
urn:nbn:se:umu:diva-124103 (URN)10.1016/j.str.2016.03.025 (DOI)000377782200011 ()27133024 (PubMedID)
External cooperation:
Available from: 2016-07-17 Created: 2016-07-17 Last updated: 2018-06-07Bibliographically approved
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