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Modeling of the elongation and retraction of Escherichia coli P pili under strain by Monte Carlo simulations
Umeå University, Faculty of Science and Technology, Applied Physics and Electronics.
Umeå University, Faculty of Science and Technology, Physics.
Umeå University, Faculty of Science and Technology, Physics.
2008 (English)In: European Biophysics Journal, ISSN 0175-7571, E-ISSN 1432-1017, Vol. 37, no 4, 381-391 p.Article in journal (Refereed) Published
Abstract [en]

P pili are fimbrial adhesion organelles expressed by uropathogenic Escherichia coli in the upper urinary tract. They constitute a stiff helix-like polymer consisting of a number of subunits joined by head-to-tail bonds. The elongation and retraction properties of individual P pili exposed to strain have been modeled by Monte Carlo (MC) simulations. The simulation model is based upon a three-state energy landscape that deforms under an applied force. Bond opening and closure are modeled by Bells theory while the elongation of the linearized part of the pilus is described by a worm-like chain model. The simulations are compared with measurements made by force measuring optical tweezers. It was found that the simulations can reproduce pili elongation as well as retraction, under both equilibrium and dynamic conditions, including entropic effects. It is shown that the simulations allow for an assessment of various model parameters, e.g. the unfolding force, energy barrier heights, and various distances in the energy landscape, including their stochastic spread that analytical models are unable to do. The results demonstrate that MC simulations are useful to model elongation and retraction properties of P pili, and therefore presumably also other types of pili, exposed to strain and/or stress. MC simulations are particularly suited for description of helix-like pili since these have an intricate self-regulating mechanical elongation behavior that makes analytical descriptions non-trivial when dynamic processes are studied, or if additional interactions in the rod or the behavior of the adhesion tip needs to be modeled.

Place, publisher, year, edition, pages
2008. Vol. 37, no 4, 381-391 p.
Keyword [en]
P pili, Escherichia coli, Optical tweezers, Monte Carlo simulations, Unfolding, Force spectroscopy
Identifiers
URN: urn:nbn:se:umu:diva-2749DOI: 10.1007/s00249-007-0223-6OAI: oai:DiVA.org:umu-2749DiVA: diva2:141008
Available from: 2007-11-08 Created: 2007-11-08 Last updated: 2017-12-14Bibliographically approved
In thesis
1. Construction of force measuring optical tweezers instrumentation and investigations of biophysical properties of bacterial adhesion organelles
Open this publication in new window or tab >>Construction of force measuring optical tweezers instrumentation and investigations of biophysical properties of bacterial adhesion organelles
2007 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Optical tweezers are a technique in which microscopic-sized particles, including living cells and bacteria, can be non-intrusively trapped with high accuracy solely using focused light. The technique has therefore become a powerful tool in the field of biophysics. Optical tweezers thereby provide outstanding manipulation possibilities of cells as well as semi-transparent materials, both non-invasively and non-destructively, in biological systems. In addition, optical tweezers can measure minute forces (< 10-12 N), probe molecular interactions and their energy landscapes, and apply both static and dynamic forces in biological systems in a controlled manner. The assessment of intermolecular forces with force measuring optical tweezers, and thereby the biomechanical structure of biological objects, has therefore considerably facilitated our understanding of interactions and structures of biological systems.

Adhesive bacterial organelles, so called pili, mediate adhesion to host cells and are therefore crucial for the initial bacterial-cell contact. Thus, they serve as an important virulence factor. The investigation of pili, both their biogenesis and their expected in vivo properties, brings information that can be of importance for the design of new drugs to prevent bacterial infections, which is crucial in the era of increased bacterial resistance towards antibiotics.

In this thesis, an experimental setup of a force measuring optical tweezers system and the results of a number of biomechanical investigations of adhesive bacterial organelles are presented. Force measuring optical tweezers have been used to characterize three different types of adhesive organelles under various conditions, P, type 1, and S pili, which all are expressed by uropathogenic Escherichia coli. A quantitative biophysical force-extension model, built upon the structure and force response, has been developed. It is found, that this model describes the biomechanical properties for all three pili in an excellent way. Various parameters in their energy landscape, e.g., bond lengths and transition barrier heights, are assessed and the difference in behavior is compared. The work has resulted in a method that in a swift way allows us to probe different types of pili with high force and high spatial resolution, which has provided an enhanced understanding of the biomechanical function of these pili.

Abstract [sv]

Optisk pincett är en teknik i vilken mikrometerstora objekt, inkluderande levande celler och bakterier, beröringsfritt kan fångas och förflyttas med hög noggrannhet enbart med hjälp av ljus. Den optiska pincetten har därmed blivit ett kraftfullt verktyg inom biofysiken, som möjliggör enastående precisions-manipulering av celler och semi-transparenta objekt. Dessutom kan denna manipulation göras intracellulärt, dvs. utan att fysiskt öppna eller penetrera cellernas membran. Den optiska pincetten kan även mäta mycket små krafter och interaktioner (< 10-12 N) samt applicera både statiska och dynamiska krafter i biologiska system med utmärkt precision. Optisk pincett är därför en utmärkt teknik för mätning av intermolekylära krafter och för bestämning av biomekaniska strukturer och dess funktioner.

Vissa typer av bakterier har specifika vidhäftningsorganeller som kallas för pili. Dessa förmedlar vidhäftningen till värdceller och är därför viktiga vid bakteriens första kontakt. En djupare förståelse av pilis uppbyggnad och biomekanik kan därmed ge information, som kan vara vital i framtagandet av nya mediciner som förhindrar bakteriella infektioner. Detta är av stor vikt i skenet av den ökande antibiotikaresistensen i vårt samhälle.

I denna avhandling presenteras konstruktionen av en experimentell uppställning av kraftmätande optiskt pincett tillsammans med resultat från biomekaniska undersökningar av vidhäftande bakteriella organeller. Kraftmätande optisk pincett har använts för att karakterisera tre olika typer av pili, P, typ 1, och S pili, vilka kan uttryckas av uropatogena Escherichia coli. En kvantitativ biofysikalisk modell som beskriver deras förlängningsegenskaper under pålagd kraft har konstruerats. Modellen bygger på pilis strukturella uppbyggnad samt på dess respons som uppmäts med den kraftmätande optiska pincetten. Modellen beskriver de biomekaniska egenskaperna väl för alla tre pili. Dessutom kan ett antal specifika bindnings- och subenhetsparametrar bestämmas, t.ex. interaktionsenergier och bindningslängder. Skillnaden mellan dessa parametrar hos de tre pilis samt deras olika kraftrespons har jämförts. Detta arbete har dels resulterat i en förbättrad förståelse av pilis biomekaniska funktion och dels i en metod som, med hög noggrannhet, tillåter oss att bestämma ett antal biomekaniska egenskaper hos olika organeller på ett effektivt sätt.

Place, publisher, year, edition, pages
Umeå: Fysik, 2007. 79 p.
Keyword
optical tweezers, biological physics, unfolding, Escherichia coli, force measurements, energy landscape, dynamic force spectroscopy, manipulation, polymers, pili
National Category
Physical Sciences
Identifiers
urn:nbn:se:umu:diva-1425 (URN)978-91-7264-435-9 (ISBN)
Public defence
2007-11-30, N450, Naturvetarhuset, Umeå Universitet Campus, Umeå, 13:00 (English)
Opponent
Supervisors
Available from: 2007-11-08 Created: 2007-11-08 Last updated: 2009-08-19Bibliographically approved
2. A study of bacterial adhesion on a single-cell level by means of force measuring optical tweezers and simulations
Open this publication in new window or tab >>A study of bacterial adhesion on a single-cell level by means of force measuring optical tweezers and simulations
2009 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The intriguing world of microbiology is nowadays accessible for detailed exploration at a single–molecular level. Optical tweezers are a novel instrument that allows for non–invasive manipulation of single cells by the sole use of laser light and operates on the nano– and micrometer scale which corresponds to the same length scale as living cells. Moreover, forces within the field of microbiology are typically in the picoNewton range which is in accordance with the capability of force measuring optical tweezers systems. Both these conformabilities imply that force measuring optical tweezers is suitable for studies of single living cells. This thesis focuses on the mechanisms of bacterial attachments to host cells which constitute the first step in bacterial infection processes. Bacteria bind specifically to host receptors by means of adhesins that are expressed either directly on the bacterial membrane or on micrometer–long adhesion organelles that are called pili. The properties of single adhesin–receptor bonds that mediate adherence of the bacterium Helicobacter pylori are first examined at various acidities. Further on, biomechanical properties of P pili expressed by Escherichia coli are presented to which computer simulations, that capture the complex kinetics of the pili structure and precisely replicate measured data, are applied. Simulations are found to be a powerful tool for investigations of adhesive attributes of binding systems and are utilized in the analyses of the specific binding properties of P pili on a single–pilus level. However, bacterial binding systems generally involve a multitude of adhesin–receptor bonds. To explore bacterial attachments, the knowledge from single–pilus studies is brought into a full multipili attachment scenario which is analyzed by means of theoretical treatments and simulations. The results are remarkable in several aspects. Not only is it found that the intrinsic properties of P pili are composed in an optimal combination to promote strong multipili bindings. The properties of the pili structure itself are also found to be optimized with respect to its in vivo environment. Indeed, the true meaning of the attributes derived at a single–pilus level cannot be unraveled until a multipili–binding system is considered. Whereas detailed studies are presented for the helix–like P pili expressed by Gram–negative Escherichia coli, conceptual studies are presented for the open coil–like T4 pili expressed by Gram–positive Streptococcus pneumoniae. The structural and adhesive properties of these two types of pili differ considerably. These dissimilarities have far–reaching consequences on the adhesion possibilities at both single–pilus and multipili levels which are discussed qualitatively. Moreover, error analyses of conventional data processing methods in dynamic force spectroscopy as well as development of novel analysis methods are presented. These findings provide better understanding of how to perform refined force measurements on single adhesion organelles as well as how to improve the analyses of measurement data to obtain accurate parameter values of biomechanical entities. In conclusion, this thesis comprises a study of bacterial adhesion organelles and the way they cooperate to establish efficient attachment systems that can successfully withstand strong external forces that acts upon bacteria. Such systems can resist, for instance, rinsing effects and thereby allow bacteria to colonize their host. By understanding the complexity, and thereby possible weaknesses, of bacterial attachments, new targets for combating bacterial infections can be identified.

Place, publisher, year, edition, pages
Umeå: Print & Media, 2009. 108 p.
National Category
Physical Sciences
Research subject
Physics; cellforskning; Molecular Cellbiology
Identifiers
urn:nbn:se:umu:diva-21493 (URN)978-91-7264-765-7 (ISBN)
Public defence
2009-05-08, N420, Naturvetarhuset, Umeå Universitet, Umeå, 10:00 (English)
Opponent
Supervisors
Available from: 2009-04-17 Created: 2009-04-14 Last updated: 2009-04-17Bibliographically approved

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