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

October 11, 2018


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


Danyelle Toyama, Mariana Abrahão Buenode Morais, Felipe Cardoso Ramos, Letícia Maria Zanphorlin, Celisa Caldana Costa Tonoli, Augusto Furio Balula, Fernando Pellonde Miranda, Vitor Medeiros Almeida, Sandro Roberto Marana, Roberto Ruller, Mario Tyago Murakami, Flavio Henrique-Silva




Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics


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 β-glucosidase (glycoside hydrolase family 1) gene from a metagenome from Lake Poraquê in the Amazon region. The gene encodes a protein of 52.9 kDa, named AmBgl-LP, which was recombinantly expressed in Escherichia coli and biochemically and structurally characterized. Although AmBgl-LP hydrolyzed the synthetic substrate p-nitrophenyl-β-d-glucopyranoside (pNPβG) and the natural substrate cellobiose, it showed higher specificity for pNPβG (kcat/Km = 6 s−1·mM−1) than cellobiose (kcat/Km = 0.6 s−1·mM−1). AmBgl-LP showed maximum activity at 40 °C and pH 6.0 when pNPβG was used as the substrate. Glucose is a competitive inhibitor of AmBgl-LP, presenting a Ki 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.




Circular dichroism, Secondary structure, Protein folding, Thermal stability, Thermodynamics, Biochemistry