Investigating mechanical properties of a protein by single molecule atomic force microscopy
department or school: Science, Faculty of
department or school: Chemistry, Department of
data provider: DSpace
degree campus: UBCV
creator: Shen, Tao
issued: 2012-05-17T16:57:30Z
issued: 2012
related degree: Master of Science - MSc
degree grantor: University of British Columbia
description: Single molecule AFM is a powerful technique affording the opportunity to understand the mechanical properties of proteins at the level of a single molecule. In Combining with the protein engineering techniques, single molecule allows us to understand protein folding/unfolding mechanisms and develop methods to tune the mechanical stability. Here, we use a small protein, GB1, the B1 IgG binding domain of protein G from Streptococcus, as a model system. In this thesis, we employed bi-His metal binding sites to probe the mechanical unfolding transition state of GB1 and rationally enhance the mechanical stability of GB1 mutant, G6-53. The transition state cannot be trapped and detected by the usual structural methods because of its high free energy. It remains a challenging task and research focus. In Chapter 3, we directly probed the mechanical unfolding transition state structure of protein GB1. The results demonstrate that the contacts between the force-bearing strands 1 and 4 are largely disrupted at the transition state, whereas the first β-hairpin and α-helix were largely intact. The second hairpin was partially disrupted. These results are in close agreement with, and provide a benchmark for, MD simulations. The mechanical stability is critical for the overall mechanical properties of elastomeric proteins. Elastomeric proteins provide tissues with extensibility, elasticity, and mechanical strength. In Chapter 4, we enhanced the mechanical stability of G6-53 with different metal ions. We demonstrated that all four divalent metal ions, Ni²⁺, Co²⁺, Zn²⁺ and Cu²⁺, enhance the mechanical stability of G6-53 to different degrees. Because this process is completely reversible, the protein can be treated like a switch. Moreover, the resultant unfolding force difference between Co²⁺ and Zn²⁺ or Zn²⁺ and Cu²⁺ is ~ 20 pN. Thus, various metal ions can be used to fine tune the mechanical stability of proteins.
aggregated CHO: https://circle.library.ubc.ca/rest/handle/2429/42335
note: Investigating mechanical properties of a protein by single molecule atomic force microscopy by TAO SHEN A THESIS SUBMITTED IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF MASTER OF SCIENCE in The Faculty of Graduate Studies (Chemistry) THE UNIVERSITY OF BRITISH COLUMBIA (Vancouver) May 2012 © Tao Shen, 2012 ii Abstract Single molecule AFM is a powerful technique affording the opportunity to understand the mechanical properties of proteins at the level of a single molecule. In Combining with the protein engineering techniques, single molecule allows us to understand protein folding/unfolding mechanisms and develop methods to tune the mechanical stability. Here, we use a small protein, GB1, the B1 IgG binding domain of protein G from Streptococcus, as a model system. In this thesis, we employed bi-His metal binding sites to probe the mechanical unfolding transition state of GB1 and rationally enhance the mechanical stability of GB1 mutant, G6-53. The transition state cannot be trapped and detected by the usual structural methods because of its high free energy. It remains a challenging task and research focus. In Chapter 3, we directly probed the mechanical unfolding transition state structure of protein GB1. The results demonstrate that the contacts between the force-bearing strands 1 and 4 are largely disrupted at the transition state, whereas the first β-hairpin and α-helix were largely intact. The second hairpin was partially disrupted. These results are in close agreement with, and provide a benchmark for, MD simulations. The mechanical stability is critical for the overall mechanical properties of elastomeric proteins. Elastomeric proteins provide tissues with extensibility, elasticity, and mechanical strength. In Chapter 4, we enhanced the mechanical stability of G6-53 with different metal ions. We demonstrated that all four divalent metal ions, Ni 2+ , Co 2+ , Zn 2+ and Cu 2+ , enhance the mechanical stability of G6-53 to different degrees. Becaus…
has type: Thesis/Dissertation
date issued: 2012-11
is shown at: 10.14288/1.0062332
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
type: Text
identifier URI: http://hdl.handle.net/2429/42335