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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.)

Conformational Changes in a Hyperthermostable Glycoside Hydrolase: Enzymatic Activity Is a Consequence of the Loop Dynamics and Protonation Balance

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Author(s):
de Oliveira, Leandro C. [1] ; da Silva, Viviam M. [2] ; Colussi, Francieli [2] ; Cabral, Aline D. [2] ; de Oliveira Neto, Mario [3] ; Squina, Fabio M. [4] ; Garcia, Wanius [2]
Total Authors: 7
Affiliation:
[1] UNESP Univ Estadual Paulista, Dept Fis, Inst Biociencias Letras & Ciencias Exatas, Sao Jose Do Rio Preto, SP - Brazil
[2] Univ Fed ABC UFABC, Ctr Ciencias Nat & Humanas, Santo Andre, SP - Brazil
[3] UNESP Univ Estadual Paulista, Inst Biociencias, Dept Fis & Biofis, Botucatu, SP - Brazil
[4] Ctr Nacl Pesquisa Energia & Mat, Lab Nacl Ciencia & Tecnol Bioetanol, Campinas, SP - Brazil
Total Affiliations: 4
Document type: Journal article
Source: PLoS One; v. 10, n. 2 FEB 27 2015.
Web of Science Citations: 16
Abstract

Endo-beta-1, 4-mannanase from Thermotoga petrophila (TpMan) is a modular hyperthermostable enzyme involved in the degradation of mannan-containing polysaccharides. The degradation of these polysaccharides represents a key step for several industrial applications. Here, as part of a continuing investigation of TpMan, the region corresponding to the GH5 domain (TpManGH5) was characterized as a function of pH and temperature. The results indicated that the enzymatic activity of the TpManGH5 is pH-dependent, with its optimum activity occurring at pH 6. At pH 8, the studies demonstrated that TpManGH5 is a molecule with a nearly spherical tightly packed core displaying negligible flexibility in solution, and with size and shape very similar to crystal structure. However, TpManGH5 experiences an increase in radius of gyration in acidic conditions suggesting expansion of the molecule. Furthermore, at acidic pH values, TpManGH5 showed a less globular shape, probably due to a loop region slightly more expanded and flexible in solution (residues Y88 to A105). In addition, molecular dynamics simulations indicated that conformational changes caused by pH variation did not change the core of the TpManGH5, which means that only the above mentioned loop region presents high degree of fluctuations. The results also suggested that conformational changes of the loop region may facilitate polysaccharide and enzyme interaction. Finally, at pH 6 the results indicated that TpManGH5 is slightly more flexible at 65 degrees C when compared to the same enzyme at 20 degrees C. The biophysical characterization presented here is well correlated with the enzymatic activity and provide new insight into the structural basis for the temperature and pH-dependent activity of the TpManGH5. Also, the data suggest a loop region that provides a starting point for a rational design of biotechnological desired features. (AU)

FAPESP's process: 12/21054-9 - Biophysical studies and of the synergistic action of thermophilics enzymes involved in the hydrolysis of mannans
Grantee:Wanius José Garcia da Silva
Support type: Regular Research Grants
FAPESP's process: 11/13242-7 - Studies of enzymatic mechanisms for the improvement of bio-fuel production
Grantee:Leandro Cristante de Oliveira
Support type: Scholarships in Brazil - Post-Doctorate
FAPESP's process: 08/58037-9 - Library generation for biomass-conversion enzymes from soil metagenome
Grantee:Fábio Márcio Squina
Support type: Program for Research on Bioenergy (BIOEN) - Young Investigators Grants
FAPESP's process: 12/03503-0 - Studies of stability, flexibility and enzymatic activity of the beta-mannanase from hyperthermophilic bacterium Thermotoga petrophila
Grantee:Viviam Moura da Silva
Support type: Scholarships in Brazil - Master