This research project is divided in two fronts: 1 - Quaternary Structure and Enzyme Kinetics in beta-Glucosidases and 2 - Structure Network and Thermostability in Proteins.Front 1 - The presence of beta-glucosidases oligomers (dimers to octamers) is already well reported in the literature. However, there is no clear view of the oligomerization effect on the enzyme kinetics. Indeed, the few reports approaching this question are based only in a single enzyme activity determination. Hence, to fill this gap we plan to isolate the monomeric and dimeric forms of the recombinant beta-glucosidase Sfbgly (5CG0) using size exclusion chromatography and perform steady-state kinetic experiments with these protein forms. The separation of these Sfbgly states will be confirmed through detection using Multi-Angle Light Scattering (MALS) and Small-Angle X-Ray Scattering (SAXS). Low temperature will be used to keep the oligomeric states unaltered during the chromatography and enzyme kinetic experiments.Front 2 - The protein tertiary structure can be represented as a network as clearly established in the literature. Interesting, the network representation shows that protein residues are in close contact with any other in the structure through a short contact pathway of at maximum 4 non-covalent contacts. Moreover, a few residues, called hubs, are essential to maintain these short contact pathways. Mutations directed to these hubs and their closest neighbors tend to reduce the protein thermal stability. Based on that, it is proposed that the hub residues mediate the distribution of the thermal energy through the protein structure. To test this hypothesis, we proposed to evaluate the order parameter S2 for each residue of the enzyme HisF at different temperatures. Following that correlations between the temperature effect on S2 and the centrality ("hubness") is a potential tool to check the role of the hub residues in affecting the thermal motions through the protein structure. (AU)
Articles published in Agência FAPESP Newsletter about the research grant:
(References retrieved automatically from Web of Science and SciELO through information on FAPESP grants and their corresponding numbers as mentioned in the publications by the authors)
ALMEIDA, VITOR MEDEIROS;
CARDOSO, MARCUS VINICIUS CANGUSSU;
MATSUYAMA, BRUNO YASUI;
FERREIRA, GLAUCIO MONTEIRO;
GOULART TROSSINI, GUSTAVO HENRIQUE;
SALINAS, ROBERTO KOPKE;
LORIA, J. PATRICK;
MARANA, SANDRO ROBERTO.
Role of a high centrality residue in protein dynamics and thermal stability.
Journal of Structural Biology,
Web of Science Citations: 0.