Investigating force-induced unfolding and folding of proteins at the single molecule level
department or school: Science, Faculty of
department or school: Chemistry, Department of
data provider: DSpace
degree campus: UBCV
creator: Wang, Han
issued: 2022-09-30T07:00:00Z
issued: 2021
related degree: Doctor of Philosophy - PhD
degree grantor: University of British Columbia
description: Understanding the molecular mechanism of protein folding has always remained a challenging problem. Extensive investigations based on ensemble methods have been carried out to shed light on the protein folding problem. Compared with ensemble-averaged measurements, single-molecule force spectroscopy (SMFS) has evolved as a powerful technique to probe mechanical unfolding/folding of proteins at the single-molecule level, which provides rich details of protein conformational changes. Optical tweezers (OT), as one representative SMFS technique, have been widely used to directly monitor a protein folding process. This thesis presents a series of studies that use OT to investigate the mechanical unfolding/folding of two distinct classes of proteins: knotted/slipknotted proteins and Ca²⁺-binding metalloproteins. First, we combined OT and steered molecular dynamics (SMD) simulations to investigate a complex slipknotted protein. When stretched from the N- and C-termini, the slipknotted structure could be completely untied via multiple unfolding pathways. Upon relaxation, this protein showed complex folding behaviors involving misfolding. Our studies demonstrate the unfolding/folding mechanisms of a slipknotted protein with a complex topological conformation. Second, we used OT to mechanically tighten, untie and retie a trefoil knotted protein. Our results revealed that protein folding with a preformed knot was fast and robust; however, the folding from the unfolded polypeptide chain without the knot conformation was significantly slower, suggesting that the knotting was the rate-limiting step for the folding of this trefoil knotted protein. Next, we investigated the mechanical unfolding/folding behaviors of a Ca²⁺-binding protein, RTX (repeats-in-toxin) block V. Our results elucidated the secretion mechanism of this RTX block V and revealed that the mechanical design can ensure an efficient translocation process. Finally, we used OT to study another Ca²⁺-binding protein, RTX…
aggregated CHO: https://circle.library.ubc.ca/rest/handle/2429/79262
note: INVESTIGATING FORCE-INDUCED UNFOLDING AND FOLDING OF PROTEINS AT THE SINGLE MOLECULE LEVEL by Han Wang B.Sc., Wuhan University, 2015 A THESIS SUBMITTED IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY in THE FACULTY OF GRADUATE AND POSTDOCTORAL STUDIES (Chemistry) THE UNIVERSITY OF BRITISH COLUMBIA (Vancouver) August 2021 © Han Wang, 2021 ii The following individuals certify that they have read, and recommend to the Faculty of Graduate and Postdoctoral Studies for acceptance, the dissertation entitled: Investigating force-induced unfolding and folding of proteins at the single molecule level submitted by Han Wang in partial fulfillment of the requirements for the degree of Doctor of Philosophy in Chemistry Examining Committee: Hongbin Li, Chemistry, UBC Supervisor Suzana K Straus, Chemistry, UBC Supervisory Committee Member Russ Algar, Chemistry, UBC University Examiner Roman Krems, Chemistry, UBC University Examiner Additional Supervisory Committee Members: Katherine S. Ryan, Chemistry, UBC Supervisory Committee Member Derek P Gates, Chemistry, UBC Supervisory Committee Member iii Abstract Understanding the molecular mechanism of protein folding has always remained a challenging problem. Extensive investigations based on ensemble methods have been carried out to shed light on the protein folding problem. Compared with ensemble-averaged measurements, single-molecule force spectroscopy (SMFS) has evolved as a powerful technique to probe mechanical unfolding/folding of proteins at the single-molecule level, which provides rich details of protein conformational changes. Optical tweezers (OT), as one representative SMFS technique, have been widely used to directly monitor a protein folding process. This thesis presents a series of studies that use OT to investigate the mechanical unfolding/folding of two distinct classes of proteins: knotted/slipknotted proteins and Ca2+-binding metalloproteins. First, we combined OT and steered molecula…
has type: Thesis/Dissertation
date issued: 2021-11
is shown at: 10.14288/1.0401374
language: eng
degree discipline: Chemistry
provider: Vancouver : University of British Columbia Library
publisher: University of British Columbia
rights: Attribution-NonCommercial-NoDerivatives 4.0 International
rights URI: http://creativecommons.org/licenses/by-nc-nd/4.0/
scholar level: Graduate
contributor: Li, Hongbin
type: Text
identifier URI: http://hdl.handle.net/2429/79262