Novel glycoside hydrolases identified by screening a Chinese Holstein dairy cow rumen-derived metagenome library

Abstract

One clone encoding glycoside hydrolases was identified through functional screening of a rumen bacterial artificial chromosome (BAC) library. Of the 68 open reading frames (ORFs) predicted, one ORF encodes a novel endo-β-1,4-xylanase with two catalytic domains of family GH43 and two cellulose-binding modules (CBMs) of family IV. Partial characterization showed that this endo-xylanase has a greater specific activity than a number of other xylanases over a wide temperature range at neutral pH and could be useful in some industrial applications.

FIG. 1.

Organization of the xylanase system. Arrows indicate the location and direction of the transcription of individual ORFs.

FIG. 2.

An alignment of the amino acid sequences of the endo-xylanases UX66a and UX66b with those in family GH43. Sequences (GenBank accession no.) are for Bacteroides thetaiotaomicron (NP812573), Bacteroides fragilis (YP101639), and Cellvibrio japonicus (YP001981313). Conserved regions are indicated as follows: *, identical in all species; :, similar in all species; and •, similarities exist. The active site and catalytic residues are indicated by ▴.

FIG. 3.

Some biochemical features of the overexpressed endo-xylanase UX66. (A) pH optima of UX66 measured at 50°C; (B) temperature optima of UX66 measured at pH 6.0 with incubation for 10 min; (C) stability of UX66 under various pH conditions measured at 50°C; (D) stability of UX66 activity at different temperatures measured at pH 6.0. The values are the average of results from three replicates.