Novel Glycoside Hydrolases Identified by Screening a Chinese Holstein Dairy Cow Rumen-Derived Metagenome Library

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.The ruminal microbes possess a repertoire of hydrolases, including glycosyl hydrolases and esterases, which mediate hydrolysis and subsequent fermentation of the diets (mainly cellulose, xylan, amylopectin, amylose, pectin) (10, 18). Thus, the ruminal microbiome has been recognized as a rich source of enzymes important not only for feed and animal industries but also for the bioenergy industry (20, 26). However, this enzyme source is largely untapped because the majority of ruminal microbes remain uncultured (17). Activity-based metagenomics enables direct identification of genes and enzymes of interest by screening metagenomic libraries for desired heterologous phenotypic traits expressed in a surrogate bacterial host. Collectively, previous studies have identified 12 esterases, 10 endoglucanases, two lipases, one cyclodextrinase, one polyphenol oxidase, and one unique multifunctional glycosyl hydrolase from ruminal metagenomic libraries (2, 6, 8-9, 21, 23). These studies provided further evidence that the ruminal microbiome contains a rich source of glycosyl hydrolases possessed by as-yet-uncultured microbes. The objective of this study was to examine a bacterial artificial chromosome (BAC) library constructed previously from the ruminal samples collected from Chinese Holstein cows for hydrolases.