9fz9

Publication Abstract from PubMed

The structural and chemical diversity of beta-glucans is reflected on the variety of essential biological roles tackled by these polysaccharides. This natural heterogeneity requires an elaborate assortment of enzymatic mechanisms to assemble, degrade or modify, as well as to extract their full biotechnological potential. Recent metagenomic efforts have provided an unprecedented growth in potential new biocatalysts, most of which remain unconfirmed or uncharacterized. Here we report the first biochemical and structural characterization of two bacterial beta-glucanases from the recently created glycoside hydrolase family 157 (LaGH157 and BcGH157) and investigate their molecular basis for substrate hydrolysis. Structural analysis by X-ray crystallography revealed that GH157 enzymes belong to clan GH-A, possessing a (beta/alpha)(8)-barrel fold catalytic domain, two beta-sandwich accessory domains and two conserved catalytic glutamates residues, with relative positions compatible with a retaining mechanism of hydrolysis. Specificity screening and enzyme kinetics suggest that the enzymes prefer mixed-linkage glucans over beta-1,3-glucans. Activity screening showed that both enzymes exhibit pH optimum at 6.5 and temperature optimum for LaGH157 and BcGH157 at 25 degrees C and 48 degrees C, respectively. Product analysis with HPAEC-PAD and LC-MS revealed that both enzymes are endo-1,3(4)-beta-glucanases, capable of cleaving beta-1,3 and beta-1,4-linked glucoses, when preceded by a beta-1,3 linkage. Moreover, BcGH157 needs a minimum of 4 subsites occupied for hydrolysis to occur, while LaGH157 only requires 3 subsites. Additionally, LaGH157 possesses exohydrolytic activity on beta-1,3 and branching beta-1,6 linkages. This unusual bifunctional endo-1,3(4)/exo-1,3-1,6 activity constitutes an expansion on our understanding of beta-glucan deconstruction, with the potential to inspire future applications.

Family GH157 enzyme exhibits broad linkage tolerance and a dual endo/exo-beta-glucanase activity on beta-glucans.,Caseiro C, McGregor NGS, Alves VD, Carvalho AL, Romao MJ, Davies GJ, Fontes CMGA, Bule P Int J Biol Macromol. 2024 Dec;282(Pt 6):137402. doi: , 10.1016/j.ijbiomac.2024.137402. Epub 2024 Nov 9. PMID:39528173^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.