A promiscuous xyloglucan xylosyltransferase 1 (OsaXXT1) from Ornithogalum saundersiae recognizes both cello-oligosaccharides and nonsaccharide lignans

Abstract

Xyloglucan xylosyltransferases (XXTs) are responsible for the biosynthesis of xyloglucans, polysaccharides widely used in drug delivery system. It is generally accepted that XXTs specifically recognize cello-oligosaccharides. The catalytic potential of XXTs towards small molecules has never been documented. Herein, a xyloglucan xylosyltransferase OsaXXT1 from Ornithogalum saundersiae was demonstrated to be reactive with small molecules. Specifically, a full-length osaxxt1 gene, which was predicted to encode a transmembrane protein, was firstly isolated from O. saundersiae based on RNA-seq data analysis. Next, the soluble expression of a truncated OsaXXT1Δ1, in which the N-terminal transmembrane region was deleted, was achieved in Escherichia coli. The soluble OsaXXT1Δ1 displayed an XXT activity, xylosylating cellohexaose to form corresponding metabolites. Subsequently, the truncated OsaXXT1Δ1 was used as the biocatalyst to react with each of 70 drug-like molecules to probe its acceptor promiscuity. Schisandrone and wulignan A1 were thus demonstrated to be reactive with UDP-xylose (UDP-Xyl) under the action of OsaXXT1Δ1. Moreover, OsaXXT1Δ1 could glucosylate schisandrone and wuligan A to form corresponding glucosides. The protein engineering towards OsaXXT1Δ1 was performed to improve its catalytic efficiency. Eighteen OsaXXT1Δ1 variants were thus generated and all these variants exhibited an improved xylosylation efficiency towards schisandrone. Taken together, this study not only provided a new knowledge for catalytic capacity of plant XXTs, but also built a new biocatalyst tool box for xylosides biosynthesis.