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(Reference retrieved automatically from Web of Science through information on FAPESP grant and its corresponding number as mentioned in the publication by the authors.)

Homodimerization of a glycoside hydrolase family GH1 beta-glucosidase suggests distinct activity of enzyme different states

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Author(s):
Otsuka, Felipe A. M. [1] ; Chagas, Rafael S. [1] ; Almeida, Vitor M. [1] ; Marana, Sandro R. [1]
Total Authors: 4
Affiliation:
[1] Univ Sao Paulo, Inst Quim, Dept Bioquim, Sao Paulo - Brazil
Total Affiliations: 1
Document type: Journal article
Source: Protein Science; v. 29, n. 9 JUL 2020.
Web of Science Citations: 0
Abstract

In this work, we investigated how activity and oligomeric state are related in a purified GH1 beta-glucosidase fromSpodoptera frugiperda(Sf beta gly). Gel filtration chromatography coupled to a multiple angle light scattering detector allowed separation of the homodimer and monomer states and determination of the dimer dissociation constant (K-D), which was in the micromolar range. Enzyme kinetic parameters showed that the dimer is on average 2.5-fold more active. Later, we evaluated the kinetics of homodimerization, scanning the changes in the Sf beta gly intrinsic fluorescence over time when the dimer dissociates into the monomer after a large dilution. We described how the rate constant of monomerization (k(off)) is affected by temperature, revealing the enthalpic and entropic contributions to the process. We also evaluated how the rate constant (k(obs)) by which equilibrium is reached after dimer dilution behaves when varying the initial Sf beta gly concentration. These data indicated that Sf beta gly dimerizes through the conformational selection mechanism, in which the monomer undergoes a conformational exchange and then binds to a similar monomer, forming a more active homodimer. Finally, we noted that conformational selection reports and experiments usually rely on a ligand whose concentration is in excess, but for homodimerization, this approach does not hold. Hence, since our approach overcomes this limitation, this study not only is a new contribution to the comprehension of GH1 beta-glucosidases, but it can also help to elucidate protein interaction pathways. (AU)

FAPESP's process: 18/18537-4 - Correlation of the quaternary structure and enzyme activity in the beta-glucosidase of Spodoptera frugiperda (Sfbgly)
Grantee:Felipe Akihiro Melo Otsuka
Support type: Scholarships in Brazil - Doctorate (Direct)
FAPESP's process: 18/25952-8 - Catalysis and Thermostability in Enzymes: Oligomerization, Structure Network and Dynamics Effects
Grantee:Sandro Roberto Marana
Support type: Regular Research Grants
FAPESP's process: 16/22365-9 - Enzyme structural network and dynamics
Grantee:Sandro Roberto Marana
Support type: Regular Research Grants