Title
BIOCHEMICAL AND BIOPHYSICAL PROPERTIES OF A METAGENOME-DERIVED GH5 ENDOGLUCANASE DISPLAYING AN UNCONVENTIONAL DOMAIN ARCHITECTURE
Author
Agnes C. Pimentel, Gabriela C.G. Ematsu, Marcelo V. Liberato, Douglas A.A. Paixão, João Paulo L.Franco Cairo, Fernanda Mandelli, Robson Tramontina, César A. Gandin, Mario de Oliveira Neto, Fabio M. Squina, Thabata M. Alvarez
Year
2017
Journal
International Journal of Biological Macromolecules
Abstract
Endoglucanases are key enzymes in the degradation of cellulose, the most abundant polymer on Earth. The aim of this work was to perform the biochemical and biophysical characterization of CelE2, a soil metagenome derived endoglucanase. CelE2 harbors a conserved domain from glycoside hydrolase family 5 (GH5) and a C-terminal domain with identity to Calx-beta domains. The recombinant CelE2 displayed preference for hydrolysis of oat beta-glucan, followed by lichenan and carboxymethyl cellulose. Optimum values of enzymatic activity were observed at 45 °C and pH 5.3, and CelE2 exhibited considerable thermal stability at 40 °C for up to 360 minutes. Regarding the cleavage pattern on polysaccharides, the release of oligosaccharides with a wide degree of polymerization indicated a characteristic of endoglucanase activity. Furthermore, the analysis of products generated from the cleavage of cellooligosaccharides suggested that CelE2 exhibited transglycosylation activity. Interestingly, the presence of CaCl2positively affect CelE2, including in the presence of surfactants. SAXS experiments provided key information on the effect of CaCl2 on the stability of CelE2 and dummy residues and rigid-body models were generated. To the best of our knowledge this is the first biochemical and biophysical characterization of an endoglucanase from family GH5 displaying this unconventional modular organization.
Instrument
J-815
Keywords
Circular dichroism, Secondary structure, Thermal stability, Thermodynamics, Biochemistry