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  • About
  • The Global ETD Search service is a free service for researchers to find electronic theses and dissertations. This service is provided by the Networked Digital Library of Theses and Dissertations.
    Our metadata is collected from universities around the world. If you manage a university/consortium/country archive and want to be added, details can be found on the NDLTD website.
11

Einzelmolekül-Kraftspektroskopie zur Untersuchung der Wechselwirkungen zwischen Tau-Peptiden und monoklonalen Antikörpern

Stangner, Tim 11 March 2015 (has links)
In dieser Dissertation werden die Bindungseigenschaften von Rezeptor-Ligand-Komplexen mit Hilfe von Optischen Pinzetten untersucht. Aufgrund ihrer außerordentlichen Orts- (2nm) und Kraftauflösung (0,2pN) ist es möglich, diese spezifischen Interaktionen anhand einzelner Bindungsereignisse zu charakterisieren. Als Modellsysteme dienen die Wechselwirkungen zwischen den phosphorylierungsspezifischen, monoklonalen Antikörpern HPT-101 und HPT-104 und dem Morbus Alzheimer relevanten Tau-Peptid. Dieses pathogen veränderte Peptid wird krankheitsspezifisch an den Aminosäuren Threonin231 und Serin235 phosphoryliert, sodass die Detektion dieses Phosphorylierungsmusters mit Hilfe von monoklonalen Antikörpern eine mögliche Früherkennung der Alzheimer-Krankheit darstellt. Eine notwendige Voraussetzung dafür ist jedoch die exakte Kenntnis der Bindungsstellen des Liganden am Rezeptor. Ziel des ersten Teils dieser Arbeit ist es, das Epitop des monoklonalen Antikörpers HPT-101 zu bestimmen. Dazu werden mögliche bindungsrelevante Aminosäuren durch ein Alanin ausgetauscht (Alanin-Scan) und so insgesamt sieben neue Tau-Isoformen aus dem ursprünglichen doppelt-phosphorylierten Peptid Tau[pThr231/pSer235] hergestellt. Die jeweiligen Interaktionen zwischen den modifizierten Peptiden und dem Antikörper werden mit der dynamischen Kraftspektroskopie untersucht und mit Hilfe eines literaturbekannten Modells analysiert. Die sich daraus ergebenden Bindungsparameter (Lebensdauer der Bindung, charakteristische Bindungslänge, freie Aktivierungsenergie und Affinitätskonstante) werden zusammen mit den relativen Bindungshäufigkeiten erstmals genutzt, um Kriterien für essentielle, sekundäre und nicht-essentielle Aminosäuren im Tau-Peptid zu definieren. Bemerkenswerterweise existieren für insgesamt drei dieser Parameter (Bindungslebensdauer, Bindungslänge und Affinitätskonstante) scharfe Klassengrenzen, mit denen eine objektive Einteilung des Epitops von Antikörper HPT-101 möglich ist. Die erhaltenen Ergebnisse sind in überzeugender Weise im Einklang mit ELISA-Messungen zu diesem Antikörper-Peptid-Komplexen, sie liefern jedoch einen tieferen Einblick in die Natur einer spezifischen Bindung, da den kraftspektroskopischen Messungen auch die Bindungskinetik zugänglich ist. Das zweite Projekt der vorliegenden Dissertation etabliert eine Methodik, um die Datenvarianz in der Bestimmung der relativen Bindungshäufigkeit zu reduzieren. Anhand einer Kombination aus Fluoreszenz- und kraftspektroskopischen Messungen werden die Wechselwirkungen zwischen dem monoklonalen Antikörper HPT-104 und dem fluoreszenzmarkierten Peptid Tau[Fl-pThr231] untersucht. Es wird gezeigt, dass durch Vorsortieren der Peptid-beschichteten Kolloide, entsprechend ihrer Oberflächenbeladung, die Datenvarianz in den Bindungshäufigkeitsmessungen signifikant reduziert wird.
12

Kinetics and dynamics of single biomolecules

Sturm, Sebastian 11 August 2016 (has links)
This thesis contains several contributions to the theoretical description and interpretation of biophysical single-molecule measurements: (i) For semiflexible polymers, we derive an efficient formulation of their local transverse dynamics in terms of a Generalized Langevin Equation. The elastic and frictional properties of the polymer are condensed into a memory kernel that is a function of the polymer\''s length and stiffness, the level of backbone tension, the position of the force probe along the polymer backbone and the boundary conditions at the polymer ends. At short times, the memory kernel attains a universal limiting form that depends neither on the polymer length nor on the boundary conditions; we obtain analytical results that accurately describe this regime. We discuss how to quickly and reliably evaluate the memory kernel for arbitrary times using a spectral decomposition method, and use an extensive body of numerical data to obtain analytical approximations to the memory kernel that cover the complementary long-time limit wherein polymer friction can be subsumed under a renormalized drag coefficient. (ii) Based on a systematic nonequilibrium treatment of an overdamped, one-dimensional stochastic escape process driven by external force, we develop a theory of Dynamic Force Spectroscopy (DFS) that generalizes previously available DFS theories to the high loading rates realized in novel experimental assays and in computer simulations. (iii) Extrapolating to future DFS experiments that may operate at far higher time resolution than presently achievable, we discuss the fast nonequilibrium relaxation of a semiflexible linker after bond rupture. Based on a rigorous theory of tension propagation in semiflexible polymers, we predict the relaxation of force within the force actuator, show that this relaxation is dominated by linker contraction, and demonstrate quantitative agreement of our predictions with experimental data obtained by a collaborating experimentalist group.
13

Observing molecular interactions that determine stability, folding, and functional states of single Na+/H+ antiporters

Kedrov, Alexej 20 November 2006 (has links)
Selective ion and solute transport across cell membranes is a vital process occurring in all types of cells. Evolutionarily developed transport proteins work as membrane-embedded molecular machines, which alternately open a gate on each side of the membrane to bind and translocate specific ions. Sodium/proton exchange plays a crucial role in maintaining cytoplasmic pH and membrane potential, while, if not regulated, the process causes severe heart diseases in humans. Here I applied single-molecule force spectroscopy to investigate molecular interactions determining the structural stability of the sodium/proton antiporter NhaA of Escherichia coli, which serves as a model system for this class of proteins. Mechanical pulling of NhaA molecules embedded in the native lipid bilayer caused a step-wise unfolding of the protein and provided insights into its stability. Modified experiments allowed observing refolding of NhaA molecules and estimating folding kinetics for individual structural elements, as well as detecting eventual misfolded conformations of the protein. The activity of NhaA increases 2000fold upon switching pH from 6 to 8. Single-molecule force measurements revealed a reversible change in molecular interactions within the ligand-binding site of the transporter at pH 5.5. The effect was enhanced in the presence of sodium ions. The observation suggests an early activation stage of the protein and provides new insights into the functioning mechanism. When studying interactions of NhaA with the inhibitor 2-aminoperimidine, I exploited single-molecule force measurements to validate the binding mechanism and to describe quantitatively formation of the protein:inhibitor complex. The ability of single-molecule force measurements to probe structurally and functionally important interactions of membrane proteins opens new prospects for using the approach in protein science and applied research.
14

Binding forces in metallo-supramolecular coordination compounds

Gensler, Manuel 15 March 2017 (has links)
Multivalente Wechselwirkungen sind in diversen biomolekularen und supramolekularen Systemen anzutreffen. Gewöhnlich werden sie durch ihre thermische Stabilität charakterisiert. Doch auch das mechanische Reißverhalten ist relevant: Ein System mit großer Reißlänge (Verformbarkeit) weist zwar eine geringere Reißkraft auf, kann aber besser auf äußere Einflüsse ohne Bindungsbruch reagieren. Daher besteht ein zunehmendes Interesse an Modellen zur Vorhersage der mechanischen Stabilität multivalenter Wechselwirkungen. Einzelmolekül-Kraftspektroskopie (SMFS) ist eine nützliche Methode, um den Reißprozess nichtkovalenter Wechselwirkungen zu studieren. Im Rahmen dieser Dissertation wurden mono- und bivalenten Pyridine, komplexiert und verbunden durch Cu(II) und Zn(II), entworfen und untersucht. Die drei bivalenten Pyridine wiesen unterschiedlich flexible Rückgratstrukturen auf (flexibel, teilflexibel, steif). Überraschenderweise wurde ein anderer Trend für die Verformbarkeiten gemessen (flexibel > steif > teilflexibel). Durch Vergleich von experimentellen Daten mit ab-initio Berechnungen konnten komplexe Reißmechanismen vorgeschlagen werden: Das Lösungsmittel war entscheidend und führte zu wasserverbrückten Zwischenprodukten, was die Verformbarkeit aller Systeme stark erhöhte. Im bivalente System mit teilflexiblem Rückgrat, koordiniert durch Cu(II), rissen beide Bindungen gleichzeitig bei vergleichsweise großen Kräften. Die beiden anderen Systeme mit Cu(II) wurden in zweistufigen Prozessen voneinander getrennt, was kleinere Reißkräfte zur Folge hatte. Insbesondere das flexible System war zwar thermisch stabiler, brach aber leichter als das monovalente System. Damit wurde zum ersten Mal der große Einfluss des Rückgrats, bei sonst gleicher Art von Wechselwirkung, auf die mechanische Stabilität bivalenter Wechselwirkungen gezeigt. Außerdem ist das entwickelte Modellsystem sehr nützlich für weiterführende Untersuchungen in biologisch relevanten wässrigen Lösungsmitteln. / Multivalent interactions are ubiquitous in biomolecular and supramolecular systems. They are commonly characterized by their thermal stability in terms of average bond lifetime or equilibration constant. However, also mechanical stabilities are relevant: A system with high rupture length (malleability) has a lower rupture force, but can more easily adopt to external constraints without rupture. Thus it is of ever-increasing interest to find appropriate models that allow predictions on the mechanical stability of multivalent interactions. Single-molecule force spectroscopy (SMFS) is a powerful tool to study the rupture process of non-covalent interactions. In the present thesis, a comprehensive study on the mechanical stability of bivalent pyridine coordination compounds with the metal ions Cu(II) and Zn(II) was performed. Surprisingly, three different backbone flexibilities (high, intermediate, low) did not correlate with the measured malleabilities (high > low > intermediate). Instead, comparison between experimental results and ab-initio calculations revealed more complex underlying rupture mechanisms: Due to the aqueous environment, hydrogen bound complexes were formed and important intermediate structures that strongly increased malleabilities. Both interactions of the intermediately flexible bivalent system with Cu(II) broke simultaneous, yielding comparatively large rupture forces. The bivalent interactions of high and low backbone flexibility with Cu(II) broke stepwise at smaller forces. Although being thermally more stable, the highly flexible system even broke at lower forces than the monovalent system. Thereby it was shown for the first time, that rupture forces of similar systems can be tuned over a broad range, just by changing the connecting backbone structure. Furthermore, the developed approach is a rich toolkit to study further the balanced interplay between rupture force and malleability in biologically relevant aqueous solvents.
15

Single-Molecule Measurements of Complex Molecular Interactions in Membrane Proteins using Atomic Force Microscopy / Einzelmolekül-Messungen komplexer molekularer Wechselwirkungen in Membranproteinen unter Benutzung des Rasterkraftmikroskops

Sapra, K. Tanuj 04 April 2007 (has links) (PDF)
Single-molecule force spectroscopy (SMFS) with atomic force microscope (AFM) has advanced our knowledge of the mechanical aspects of biological processes, and helped us take big strides in the hitherto unexplored areas of protein (un)folding. One such virgin land is that of membrane proteins, where the advent of AFM has not only helped to visualize the difficult to crystallize membrane proteins at the single-molecule level, but also given a new perspective in the understanding of the interplay of molecular interactions involved in the construction of these molecules. My PhD work was tightly focused on exploiting this sensitive technique to decipher the intra- and intermolecular interactions in membrane proteins, using bacteriorhodopsin and bovine rhodopsin as model systems. Using single-molecule unfolding measurements on different bacteriorhodopsin oligomeric assemblies - trimeric, dimeric and monomeric - it was possible to elucidate the contribution of intra- and interhelical interactions in single bacteriorhodopsin molecules. Besides, intriguing insights were obtained into the organization of bacteriorhodopsin as trimers, as deduced from the unfolding pathways of the proteins from different assemblies. Though the unfolding pathways of bacteriorhodopsin from all the assemblies remained the same, the different occurrence probability of these pathways suggested a kinetic stabilization of bacteriorhodopsin from a trimer compared to that existing as a monomer. Unraveling the knot of a complex G-protein coupled receptor, rhodopsin, showed the existence of two structural states, a native, functional state, and a non-native, non-functional state, corresponding to the presence or absence of a highly conserved disulfide bridge, respectively. The molecular interactions in absence of the native disulfide bridge mapped onto the three-dimensional structure of native rhodopsin gave insights into the molecular origin of the neurodegenerative disease retinitis pigmentosa. This presents a novel technique to decipher molecular interactions of a different conformational state of the same molecule in the absence of a high-resolution X-ray crystal structure. Interestingly, the presence of ZnCl2 maintained the integrity of the disulfide bridge and the nature of unfolding intermediates. Moreover, the increased mechanical and thermodynamic stability of rhodopsin with bound zinc ions suggested a plausible role for the bivalent ion in rhodopsin dimerization and consequently signal transduction. Last but not the least, I decided to dig into the mysteries of the real mechanisms of mechanical unfolding with the help of well-chosen single point mutations in bacteriorhodopsin. The monumental work has helped me to solve some key questions regarding the nature of mechanical barriers that constitute the intermediates in the unfolding process. Of particular interest is the determination of altered occurrence probabilities of unfolding pathways in an energy landscape and their correlation to the intramolecular interactions with the help of bioinformatics tools. The kind of work presented here, in my opinion, will not only help us to understand the basic principles of membrane protein (un)folding, but also to manipulate and tune energy landscapes with the help of small molecules, proteins, or mutations, thus opening up new vistas in medicine and pharmacology. It is just a matter of a lot of hard work, some time, and a little bit of luck till we understand the key elements of membrane protein (un)folding and use it to our advantage.
16

Single-Molecule Measurements of Complex Molecular Interactions in Membrane Proteins using Atomic Force Microscopy

Sapra, K. Tanuj 01 March 2007 (has links)
Single-molecule force spectroscopy (SMFS) with atomic force microscope (AFM) has advanced our knowledge of the mechanical aspects of biological processes, and helped us take big strides in the hitherto unexplored areas of protein (un)folding. One such virgin land is that of membrane proteins, where the advent of AFM has not only helped to visualize the difficult to crystallize membrane proteins at the single-molecule level, but also given a new perspective in the understanding of the interplay of molecular interactions involved in the construction of these molecules. My PhD work was tightly focused on exploiting this sensitive technique to decipher the intra- and intermolecular interactions in membrane proteins, using bacteriorhodopsin and bovine rhodopsin as model systems. Using single-molecule unfolding measurements on different bacteriorhodopsin oligomeric assemblies - trimeric, dimeric and monomeric - it was possible to elucidate the contribution of intra- and interhelical interactions in single bacteriorhodopsin molecules. Besides, intriguing insights were obtained into the organization of bacteriorhodopsin as trimers, as deduced from the unfolding pathways of the proteins from different assemblies. Though the unfolding pathways of bacteriorhodopsin from all the assemblies remained the same, the different occurrence probability of these pathways suggested a kinetic stabilization of bacteriorhodopsin from a trimer compared to that existing as a monomer. Unraveling the knot of a complex G-protein coupled receptor, rhodopsin, showed the existence of two structural states, a native, functional state, and a non-native, non-functional state, corresponding to the presence or absence of a highly conserved disulfide bridge, respectively. The molecular interactions in absence of the native disulfide bridge mapped onto the three-dimensional structure of native rhodopsin gave insights into the molecular origin of the neurodegenerative disease retinitis pigmentosa. This presents a novel technique to decipher molecular interactions of a different conformational state of the same molecule in the absence of a high-resolution X-ray crystal structure. Interestingly, the presence of ZnCl2 maintained the integrity of the disulfide bridge and the nature of unfolding intermediates. Moreover, the increased mechanical and thermodynamic stability of rhodopsin with bound zinc ions suggested a plausible role for the bivalent ion in rhodopsin dimerization and consequently signal transduction. Last but not the least, I decided to dig into the mysteries of the real mechanisms of mechanical unfolding with the help of well-chosen single point mutations in bacteriorhodopsin. The monumental work has helped me to solve some key questions regarding the nature of mechanical barriers that constitute the intermediates in the unfolding process. Of particular interest is the determination of altered occurrence probabilities of unfolding pathways in an energy landscape and their correlation to the intramolecular interactions with the help of bioinformatics tools. The kind of work presented here, in my opinion, will not only help us to understand the basic principles of membrane protein (un)folding, but also to manipulate and tune energy landscapes with the help of small molecules, proteins, or mutations, thus opening up new vistas in medicine and pharmacology. It is just a matter of a lot of hard work, some time, and a little bit of luck till we understand the key elements of membrane protein (un)folding and use it to our advantage.
17

Multivalency in the interaction of biological polymers

Reiter-Scherer, Valentin D. 14 September 2020 (has links)
Diese Dissertation konzentriert sich auf die Untersuchung multivalenter Wechselwirkungen zwischen Hämagglutinin (HA) sowie Neuraminidase (NA) zweier Stämme des Influenzavirus (H1N1 und H3N2) und dem zellulären Liganden Sialinsäure (SA) unter Verwendung von Rasterkraftmikroskopie und Einzelmolekülkraftspektroskopie (SMFS). Bindungskräfte sowie Dissoziations- und Assoziationskinetiken, zusammen mit den intermolekularen Potentiallandschaften wurden, nach bestem Wissen erstmalig, auf Einzelmolekülebene mittels SMFS quantifiziert. Zu diesem Zweck wurden mono- und multivalente SA-Liganden (SAPEGLA und dPGSA) eingesetzt. Abweichungen der experimentellen Kraftspektren vom klassischen Kramers-Bell-Evans-Modell vorhergesagten Verhalten wurden durch das Friddle-Noy-De Yoreo-Model berücksichtigt. NA beider Virusstämme zeigte trotz ähnlicher Bindungskräfte eine stabilere Bindung mit SA als HA und dissoziierte 3 – 7 mal langsamer. Es wird vermutet, dass die höhere Stabilität die geringere Oberflächendichte von NA auf der Virushülle im Vergleich zu HA ausgleicht. Die Bindungskräfte eines SAPEGLA-Clusters nehmen mit der Anzahl der Bindungen und die Dissoziationskinetik folgt dem theoretisch vorhergesagten Trend. Die Dissoziationsrate von NA ist etwa 6-mal höher ist als ihre katalytische Rate, weshalb Mehrfachbindungen zur Spaltung von SA erforderlich sind. Die Dissoziationsrate von N1 in der gleichen Größenordnung wie die von H3 und es wird vermutet, dass derartige Ähnlichkeiten die Übertragbarkeit des Virus begünstigen. Darüber hinaus wird gezeigt, dass die thermische Stabilität von HA-dPGSA höher ist als von HA-SAPEGLA und im Bereich von 3 - 4 Einzelbindungen liegt, was für NA-dPGSA nicht beobachtet werden konnte. Daher bindet dPGSA spezifisch und kooperativ multivalent an HA. Kompetitive Bindungstests zeigen, dass SMFS zum Screening von antiviralen Inhibitoren verwendet werden und Zugang zu deren Design auf Einzelmolekülebene liefern könnte. / This thesis focuses on studying multivalent interactions between influenza virus hemagglutinin (HA) as well as neuraminidase (NA) of two viral strains (H1N1 and H3N2) and the cellular ligand sialic acid (SA) by using scanning force microscopy and single molecule force spectroscopy (SMFS). Unbinding forces as well as dissociation and association kinetics together with the free energy landscapes were, to the best knowledge for the first time, individually quantified on the single molecule level using SMFS. To this extent, designed synthetic monovalent (SAPEGLA) and multivalent (dPGSA) SA displaying ligands were employed. Surprisingly, the experimental force spectra did not show the log-linear trend predicted by the classical Kramers-Bell-Evans model, but rather follow the more recent Friddle-Noy-De Yoreo model. NA of both viral strains forms a more stable bond with SA than HA, and dissociates 3 to 7 times slower. It is reasoned that the higher stability compensates for the lesser amount of NA compared to HA that is typically found on the viral envelope. The unbinding forces of the cluster of SAPEGLA increased gradually with the number of bonds in the cluster and the dissociation kinetics follow the theoretically predicted trend. The dissociation rate of NA was found to be about 6 times higher than its catalytic rate, indicating that multiple bonds are needed for cleavage of SA. The dissociation rate of N1 is on the same order as that of H3, suggesting that these similarities between the two strains favor transmissibility. The thermal stability of the HA-dPGSA bond is higher than the HA-SAPEGLA reaching that of three to four single bonds, proving specificity and cooperativity. Such an enhancement could not be observed for the binding of NA. This thesis also shows that SMFS could be used as a tool to screen antiviral inhibitors in competitive binding assays, which may contribute insight into the design of antiviral inhibitors on the single molecule level.
18

Structural stability of the integron synaptic complex

Vorobevskaia, Ekaterina 03 May 2024 (has links)
The predominant tool for adaptation in Gram-negative bacteria is a genetic system called integron. It rearranges gene cassettes, promoting multiple antibiotic resistances, a recognized major global health threat. It is based on a unique recombination process involving a Tyrosine recombinase – called integrase IntI – and folded single-stranded DNA hairpins – called attC sites. Four recombinases and two attC sites form a macromolecular synaptic complex, which is key to the entire recombination process and the focus of our study. The bottom strand of all attC sites shows highest recombination in vivo, however, it still varies greatly and the underlying reason is unknown. We hypothesize that the difference in recombination efficiency arises from the variable mechanical stability of the synaptic complex, which in turn is affected by the attC site. Here, we established an optical tweezers force-spectroscopy assay that allows us to probe the synaptic complex stability for different DNA substrates and protein variants. We discovered a strong correlation between recombination efficiency and the mechanical stability of the synapse, indicating a regulatory mechanism from the DNA sequence to the quaternary complex structure stability. We have discovered protein residues interacting with the DNA in trans, within the synaptic complex, which reduces its stability. Furthermore, we discovered that the C-terminal helix, a conserved structural feature of tyrosine recombinases plays a key role in the stabilization of the tetramer assembly on the DNA, which upon mutation significantly destabilized the synaptic complex. Expanding upon this new understanding of synapse stability regulation we developed a novel approach for destabilizing the synaptic complex, potentially reducing the recombination efficiency. We designed α-helix mimicking peptides that would compete with the C-terminal tail of the integrase, block the interlocking interaction, and lead to synaptic complex destabilization. We have observed a prominent destabilizing effect on the synaptic complex already at 10 µM peptide concentration. Overall, our findings reveal new regulatory mechanisms in the recombination efficiency of the bacterial integron and provide first data for the active synapse destabilization mechanism. This novel understanding of the regulatory role the synaptic complex plays in the recombination efficiency of the integron system introduces a new approach to reduce the spread of antibiotic resistance among bacteria. / Das vorherrschende Anpassungsmittel bei gramnegativen Bakterien ist ein genetisches System, das Integron genannt wird. Es ordnet Genkassetten neu an und fördert so multiple Antibiotikaresistenzen, die eine globale Gesundheitsbedrohung darstellen. Es basiert auf einem einzigartigen Rekombinationsprozess, an dem eine Tyrosin-Rekombinase - Integrase IntI genannt - und gefaltete einzelsträngige DNA-Hairpins - attC-Stellen genannt - beteiligt sind. Vier Rekombinasen und zwei attC-Stellen bilden einen makromolekularen synaptischen Komplex, der für den gesamten Rekombinationsprozess entscheidend ist und im Mittelpunkt unserer Forschung steht. Der untere Strang aller attC-Stellen weist in vivo die höchste Rekombinationsrate auf, die jedoch aus unbekannten Grund stark variier. Wir vermuten, dass der Unterschied in der Rekombinationsrate auf die unterschiedliche mechanische Stabilität des synaptischen Komplexes zurückzuführen ist, die wiederum von der attC-Stelle beeinflusst wird. Hier haben wir einen Test mittels Kraft-spektroskopie mit einer optischen Pinzette entwickelt, mit dem wir die Stabilität des synaptischen Komplexes für verschiedene DNA-Substrate und Proteinvarianten untersuchen können. Wir stellten eine starke Korrelation zwischen der Rekombinationsrate und der mechanischen Stabilität der Synapse fest, was auf einen Regulationsmechanismus zwischen der DNA-Sequenz und der Stabilität der quaternären Komplexstruktur hinweist. Wir haben Proteinreste entdeckt, die innerhalb des synaptischen Komplexes mit der DNA in trans interagieren, was zu einer Verringerung dessen Stabilität führt. Darüber hinaus stellten wir fest, dass die C-terminale Helix, ein konserviertes Strukturmerkmal von Tyrosin-Rekombinasen, eine Schlüsselrolle bei der Stabilisierung des Tetramer-Aufbaus an der DNA spielt, die bei Mutation den synaptischen Komplex erheblich destabilisiert. Auf der Grundlage dieses neuen Verständnisses der Regulierung der Synapsenstabilität haben wir einen neuen Ansatz zur Destabilisierung des synaptischen Komplexes entwickelt, der die Effizienz der Rekombination verringern könnte. Wir entwarfen α-Helix-nachahmende Peptide, die mit dem C-terminalen Ende der Integrase konkurrieren, die Interlocking-Interaktion blockieren und zur Destabilisierung des synaptischen Komplexes führen. Wir haben eine deutliche destabilisierende Wirkung auf den synaptischen Komplex bereits bei einer Peptidkonzentration von 10 µM beobachtet. Insgesamt zeigen unsere Ergebnisse neue Regulationsmechanismen für die Rekombinationsleistung des bakteriellen Integrons auf und liefern erste Daten für den Mechanismus der aktiven Destabilisierung der Synapse. Dieses neue Verständnis der regulatorischen Rolle, die der synaptische Komplex bei der Rekombinationseffizienz des Integronsystems spielt, eröffnet einen neuen Ansatz zur Verringerung der Verbreitung von Antibiotikaresistenzen unter Bakterien.

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