Attachment of sugars to nitrogen and oxygen in peptides is ubiquitous in biology, but glycosylation of sulfur atoms has only been recently described. Here, we characterize two S-glycosyltransferases SunS and ThuS that selectively glycosylate one of five Cys residues in their substrate peptides; substitution of this Cys with Ser results in a strong decrease in glycosylation activity. Crystal structures of SunS and ThuS in complex with UDP-glucose or a derivative reveal an unusual architecture in which a glycosyltransferase type A (GTA) fold is decorated with additional domains to support homodimerization. Dimer formation creates an extended cavity for the substrate peptide, drawing functional analogy with O-glycosyltransferases involved in cell wall biosynthesis. This extended cavity contains a sharp bend that may explain the site selectivity of the glycosylation because the target Cys is in a Gly-rich stretch that can accommodate the bend. These studies establish a molecular framework for understanding the unusual S-glycosyltransferases.

糖与肽中氮和氧的结合在生物学中普遍存在，但硫原子的糖基化直到最近才被描述。在这里，我们表征了两种S -糖基转移酶 SunS 和 ThuS，它们选择性地糖基化其底物肽中五个 Cys 残基之一；用 Ser 取代此 Cys 会导致糖基化活性的强烈降低。与 UDP-葡萄糖或衍生物复合的 SunS 和 ThuS 的晶体结构揭示了一种不寻常的结构，其中 A 型糖基转移酶 (GTA) 折叠装饰有额外的结构域以支持同二聚化。二聚体的形成为底物肽创造了一个扩展的空腔，与O进行功能类比-参与细胞壁生物合成的糖基转移酶。这个扩展的空腔包含一个尖锐的弯曲，这可以解释糖基化的位点选择性，因为目标 Cys 处于可以适应弯曲的富含甘氨酸的伸展中。这些研究建立了一个分子框架来理解不寻常的S-糖基转移酶。