Statistical Study of the Unfolding of Multimodular Proteins and their Energy Landscape by Atomic Force Microscopy

摘要: The aim of the present thesis is to investigate several aspects of: proteins mechanics, interprotein interactions and study also new techniques, theoretical technical, obtain analyze force spectroscopy experiments. first section dedicated statistical properties unfolding forces in a chain homomeric multimodular proteins. basic idea this kind statistic divide peaks observed extension curve separate groups then these considering their position curves. In fact protein related number not yet unfolded modules (we call it "N"). Such effect yields linear dependence most probable peak on ln(N). We demonstrate how such can be used extract kinetic parameters how, ignoring it, could lead significant errors. Following topic we continue with non methods that, using resampling from rupture any peak, reconstruct for all other chain. Then discussion about Monte Carlo simulation pulling present. framework methodology has introduced understand various simulations done. chapter introduce ligand receptor when directly functionalize AFM tip substrate. fact, many our experiments, see "cloud points" vs loading rate graph. have modeled system composed by "N" parallel springs, studying distribution obtained graph establish procedure restore used. been discuss real experiments similar biotin-avidin interaction. following order approximation Bell-Evans model where more explicit form potential considered. particular curvature applied at minimum metastable state well known prefactors transition are fixed force, however what happen nature, (that second local derivative interacting energy respect reaction coordinate its maximum) depend applied. results Laminin-binding-protein discussed, showed phase pH was changed. behavior changes, normal WLC plateau behavior. analysis curves shows length maximum prominent correspond full protein. However, that associated dimers trymers distribution. Later approach lock key mechanism, "handles" specific pattern, introduced. handles (I27)3 (I27–SNase)3 were biochemically attached to: strept-actin molecules, biotin RNase Angiogenin. main "handle-(molecule A)-(molecule B)-handle" covalently respective molecules two "A B" secondary bonds. This allows better recognition protein-protein interaction enabling us filter out spurious events. Doing comparing detachments bare handles, able information between molecule A B. last chapters preliminary character previous part thesis. estimation effective mass viscous drag cantilevers studied autocorrelation noise power spectrum. Usually spectrum method used, should give approximately same information. important high frequency modulation techniques. they needed interpret results. show good agreement cantilever Afterwards low base. allow record amplitude shift signal Using channel managed static lower level noise. Moreover signals direct dynamic stiffness lose system, standard technique would difficult (or even impossible) obtain.