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(Reference retrieved automatically from Web of Science through information on FAPESP grant and its corresponding number as mentioned in the publication by the authors.)

Spatially remote motifs cooperatively affect substrate preference of a ruminal GH26-type endo-?-1,4-mannanase

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Author(s):
Mandelli, Fernanda [1] ; de Morais, Mariana Abrahao Bueno [1] ; de Lima, Evandro Antonio [1] ; Oliveira, Leane [2] ; Persinoti, Gabriela Felix [1] ; Murakami, Mario Tyago [1]
Total Authors: 6
Affiliation:
[1] Brazilian Ctr Res Energy & Mat CNPEM, Brazilian Biorenewables Natl Lab LNBR, BR-13083970 Campinas, SP - Brazil
[2] Elanco Anim Hlth, Greenfield, IN 46140 - USA
Total Affiliations: 2
Document type: Journal article
Source: Journal of Biological Chemistry; v. 295, n. 15, p. 5012-5021, APR 10 2020.
Web of Science Citations: 0
Abstract

?-Mannanases from the glycoside hydrolase 26 (GH26) family are retaining hydrolases that are active on complex heteromannans and whose genes are abundant in rumen metagenomes and metatranscriptomes. These enzymes can exhibit distinct modes of substrate recognition and are often fused to carbohydrate-binding modules (CBMs), resulting in a molecular puzzle of mechanisms governing substrate preference and mode of action that has not yet been pieced together. In this study, we recovered a novel GH26 enzyme with a CBM35 module linked to its N terminus (CrMan26) from a cattle rumen metatranscriptome. CrMan26 exhibited a preference for galactomannan as substrate and the crystal structure of the full-length protein at 1.85 ? resolution revealed a unique orientation of the ancillary domain relative to the catalytic interface, strategically positioning a surface aromatic cluster of the ancillary domain as an extension of the substrate-binding cleft, contributing to galactomannan preference. Moreover, systematic investigation of nonconserved residues in the catalytic interface unveiled that residues Tyr(195) (?3 subsite) and Trp(234) (?5 subsite) from distal negative subsites have a key role in galactomannan preference. These results indicate a novel and complex mechanism for substrate recognition involving spatially remote motifs, distal negative subsites from the catalytic domain, and a surface-associated aromatic cluster from the ancillary domain. These findings expand our molecular understanding of the mechanisms of substrate binding and recognition in the GH26 family and shed light on how some CBMs and their respective orientation can contribute to substrate preference. (AU)

FAPESP's process: 16/19995-0 - Analysis of structural and functional diversity of GH43 enzymes from Xanthomonas axonopodis pv. citri: biological implications and potential biotechnological applications
Grantee:Mariana Abrahão Bueno de Morais
Support type: Scholarships in Brazil - Post-Doctorate
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 type: Research Projects - Thematic Grants