Modular Hyperthermostable Bacterial Endo-beta-1, 4-Mannanase: Molecular Shape, Flexibility and Temperature-Dependent Conformational Changes

Author(s)

da Silva, Viviam M. ^[1] ; Colussi, Francieli ^[1] ; Neto, Mario de Oliveira ^[2] ; Braz, Antonio S. K. ^[1] ; Squina, Fabio M. ^[3] ; Oliveira, Cristiano L. P. ^[4] ; Garcia, Wanius ^[1]

Affiliation

^[1] Univ Fed ABC UFABC, Ctr Ciencias Nat & Humanas, Sao Paulo - Brazil

^[2] Univ Estadual Paulista, Inst Biociencias, Dept Fis & Biofis, Sao Paulo - Brazil

^[3] Ctr Nacl Pesquisa Energia & Mat, Lab Nacl Ciencia & Tecnol Bioetanol, Sao Paulo - Brazil

^[4] Univ Sao Paulo, Inst Fis, BR-01498 Sao Paulo - Brazil

• Document type: Journal article
• Source: PLoS One; v. 9, n. 3 MAR 26 2014.
• Web of Science Citations: 3

Abstract

Endo-beta-1,4-mannanase from Thermotoga petrophila (TpMan) is a hyperthermostable enzyme that catalyzes the hydrolysis of beta-1,4-mannoside linkages in various mannan-containing polysaccharides. A recent study reported that TpMan is composed of a GH5 catalytic domain joined by a linker to a carbohydrate-binding domain. However, at this moment, there is no three-dimensional structure determined for TpMan. Little is known about the conformation of the TpMan as well as the role of the length and flexibility of the linker on the spatial arrangement of the constitutive domains. In this study, we report the first structural characterization of the entire TpMan by small-angle X-ray scattering combined with the three-dimensional structures of the individual domains in order to shed light on the low-resolution model, overall dimensions, and flexibility of this modular enzyme at different temperatures. The results are consistent with a linker with a compact structure and that occupies a small volume with respect to its large number of amino acids. Furthermore, at 20 degrees C the results are consistent with a model where TpMan is a molecule composed of three distinct domains and that presents some level of molecular flexibility in solution. Even though the full enzyme has some degree of molecular flexibility, there might be a preferable conformation, which could be described by the rigid-body modeling procedure. Finally, the results indicate that TpMan undergoes a temperature-driven transition between conformational states without a significant disruption of its secondary structure. Our results suggest that the linker can optimize the geometry between the other two domains with respect to the substrate at high temperatures. These studies should provide a useful basis for future biophysical studies of entire TpMan. (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 Opportunities: Regular Research 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 Opportunities: Scholarships in Brazil - Master