A novel beta-glucosidase isolated from the microbial metagenome of Lake Poraque (Amazon, Brazil)

Author(s):Show less -

Toyama, Danyelle ^[1] ; Bueno de Morais, Mariana Abrahao ^[2] ; Ramos, Felipe Cardoso ^[2] ; Zanphorlin, Leticia Maria ^[2] ; Costa Tonoli, Celisa Caldana ^[3] ; Balula, Augusto Furio ^[1] ; de Miranda, Fernando Felton ^[4] ; Almeida, Vitor Medeiros ^[5] ; Marana, Sandro Roberto ^[5] ; Ruller, Roberto ^[2] ; Murakami, Mario Tyago ^[2] ; Henrique-Silva, Flavio ^[1]

Affiliation

^[1] Univ Fed Sao Carlos, Dept Genet & Evolut, Lab Mol Biol, Rodovia Washington Luiz, Km 235, Sao Carlos, SP - Brazil

^[2] Natl Ctr Res Energy & Mat, Brazilian Bioethanol Sci & Technol Lab, Giuseppe Maximo Scolfaro 10000, Campinas, SP - Brazil

^[3] Natl Ctr Res Energy & Mat, Brazilian Biosci Natl Lab, Campinas, SP - Brazil

^[4] Petr Brasileiro SA Petrobras, Ctr Pesquisas & Desenvolvimento Leopoldo Americo, Rio De Janeiro, RJ - Brazil

^[5] Inst Quim, Dept Bioquim, Sao Paulo, SP - Brazil

• Document type: Journal article
• Source: BIOCHIMICA ET BIOPHYSICA ACTA-PROTEINS AND PROTEOMICS; v. 1866, n. 4, p. 569-579, APR 2018.
• Web of Science Citations: 5

Abstract

The Amazon region holds most of the biological richness of Brazil. Despite their ecological and biotechnological importance, studies related to microorganisms from this region are limited. Metagenomics leads to exciting discoveries, mainly regarding non-cultivable microorganisms. Herein, we report the discovery of a novel beta-glucosidase (glycoside hydrolase family 1) gene from a metagenome from Lake Poraque in the Amazon region. The gene encodes a protein of 52.9 kDa, named AmBgl-LP, which was recombinantly expressed in Escherichia soli and biochemically and structurally characterized. Although AmBgl-LP hydrolyzed the synthetic substrate pnitrophenyl-beta-D-glucopyranoside (pNP beta G) and the natural substrate cellobiose, it showed higher specificity for pNP beta G (k(cat)/K-m = 6 s(-1).mM(-1)) than cellobiose (k(cat)/K-m = 0.6 s(-1).mM(-1)) AmBgl-LP showed maximum activity at 40 degrees C and pH 6.0 when pNP beta G was used as the substrate. Glucose is a competitive inhibitor of AmBgl-LP, presenting a K-i of 14 mM. X-ray crystallography and Small Angle X-ray Scattering were used to determine the AmBgl-LP three-dimensional structure and its oligomeric state. Interestingly, despite sharing similar active site architecture with other structurally characterized GH1 family members which are monomeric, AmBgl-LP forms stable dimers in solution. The identification of new GH1 members by metagenomics might extend our understanding of the molecular mechanisms and diversity of these enzymes, besides enabling us to survey their industrial applications. (AU)

• FAPESP's process: 15/26982-0 - Exploring novel strategies for depolymerization of plant cell-wall polysaccharides: from structure, function and rational design of glycosyl hydrolases to biological implications and potential biotechnological applications
• Grantee: Mário Tyago Murakami
• Support Opportunities: Research Projects - Thematic Grants