Structural dissection of sterol glycosyltransferase UGT51 from Saccharomyces cerevisiae for substrate specificity.

Graphical abstract Highlights • UGT51 catalyzes the transformation of glucose into varied sterols. • Complexes with donor substrate UDPG was solved and highlight key residues. • Molecular docking of cholesterol into UGT51 highlight key residues. • Comparison of UGT51 with M7_1 model revealed key factors for substrate specificity. Abstract Sterol glycosyltransferases catalyze the formation of a variety of glycosylated sterol derivatives and are involved in producing a plethora of bioactive natural products. To understand the molecular mechanism of sterol glycosyltransferases, we determined crystal structures of a sterol glycosyltransferase UGT51 from Saccharomyces cerevisiae. The structures of the UGT51 and its complex with uridine diphosphate glucose (UDPG) were solved at resolutions of 2.77 Å and 1.9 Å, respectively. The structural analysis revealed that a long hydrophobic cavity, 9.2 Å in width and 17.6 Å in length located at the N-terminal domain of UGT51, is suitable for the accommodation of sterol acceptor substrates. Furthermore, a short, conserved sequence of S847-M851 was identified at the bottom of the hydrophobic cavity, which might be the steroid binding site and play an important role for the UGT51 catalytic specificity towards sterols. Molecular docking simulations indicated that changed unique interaction network in mutant M7_1 (S801A/L802A/V804A/K812A/E816K/S849A/N892D), with an 1800-fold activity improvement toward an unnatural substrate protopanaxadiol (PPD), might influence its substrate preference. This study reported the first sterol glycosyltransferase structure, providing a molecular blueprint for generating tailored sterol glycosyltransferases as potential catalytic elements in synthetic biology. [ABSTRACT FROM AUTHOR]