Synaptic vesicles studied by dynamic light scattering

Abstract

Abstract. The size polydispersity distribution of synaptic vesicles (SVs) is characterized under quasiphysiological conditions by dynamic light scattering (DLS). Highly purified fractions of SVs obtained from rat brain still contain a small amount of larger contaminant structures, which can be quantified by DLS and further reduced by asymmetric-flow field-flow (AFFF) fractionation. The intensity autocorrelation functions g 2(τ ) recorded from these samples are analyzed by a constrained regularization method as well as by an alternative direct modeling approach. The results are in quantitative agreement with the polydispersity obtained from cryogenic electron microscopy of vitrified SVs. Next, different vesicle fusion assays based on samples composed of SVs and small unilamellar proteoliposomes with the fusion proteins syntaxin 1 and SNAP-25A are characterized by DLS. The size increase of the proteoliposomes due to SNARE-dependent fusion with SVs is quantified by DLS under quasi-physiological conditions.

Synaptic vesicles studied by small-angle x-ray scattering

“…We find that DLS autocorrelation curves can consistently be described by a model calculation employing a size distribution function p n (R) obtained by cryo-EM of vitrified SV dispersions (16).…”

“…The size distribution of the native SV preparation contains a second trace population of larger particles (14,15). Asymmetric-flow field-flow (AFFF) fractionation is used for further purification of the SV samples, effectively eliminating the larger contaminant particles (16). DLS spectra of SV populations can be regularized and inverted, giving di-…”

“…Entropic contributions to the free energy due to protein cluster formation and disintegration on the SV is investigated by calculating the entropy of a microcanonical SV model. Chapter 6 presents work on a cell free fusion system, employing SVs and proteo-liposomes with reconstituted SNARE proteins (16). The capabilities of using DLS to quantify fusion processes is assessed (16).…”

“…Chapter 6 presents work on a cell free fusion system, employing SVs and proteo-liposomes with reconstituted SNARE proteins (16). The capabilities of using DLS to quantify fusion processes is assessed (16). A SAXS Gedankenexperiment is considered, and calculated scattering curves of fused SVs and proteo-liposomes are presented and discussed (18).…”

“…A SAXS Gedankenexperiment is considered, and calculated scattering curves of fused SVs and proteo-liposomes are presented and discussed (18). Chapter 7 finally summarizes the results and presents the conclusions (15)(16)(17)(18).…”