Martini Rachel M, van der Donk Wilfred A
Department of Biochemistry University of Illinois at Urbana-Champaign, Urbana, Illinois 61801, USA.
Department of Chemistry and Howard Hughes Medical Institute, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801, USA.
bioRxiv. 2025 May 20:2025.05.19.655019. doi: 10.1101/2025.05.19.655019.
Glycocins are a growing family of ribosomally synthesized and posttranslationally modified peptides that are - and/or -glycosylated. Using a sequence similarity network of putative glycosyltransferases, the biosynthetic gene cluster was identified in the genome of . ThtA is the precursor peptide to a member of the glycocin F family of glycocins. Like other members of this family, the glycosyltransferase (ThtS) encoded in the biosynthetic gene cluster adds -acetyl-glucosamine to both Ser and Cys residues of ThtA. -linked glycosylation has been shown to be chemically and enzymatically resistant to cleavage and therefore ThtS may be a valuable starting point for engineering efforts. The glycocin derived from ThtA, which we name thermoglycocin, was structurally characterized. Thermoglycocin is unique in that in addition to two nested disulfide bonds, it contains an additional disulfide bond creating a C-terminal loop. Unexpectedly, ThtA lacks the common double glycine motif that denotes a C39-peptidase leader peptide cleavage site. Based on AlphaFold3 modeling, we postulate that cleavage between the leader and core peptide occurs instead at a GK motif. This study adds to the small number of characterized glycocins, employs AlphaFold3 to aid in predicting the structure of the mature peptide product, and suggests a common naming convention similar to that established for lanthipeptides.
糖菌素是一类不断增加的核糖体合成且经过翻译后修饰的肽，这些肽被进行了O-和/或N-糖基化。利用假定糖基转移酶的序列相似性网络，在……的基因组中鉴定出了生物合成基因簇。ThtA是糖菌素F家族中一种糖菌素的前体肽。与该家族的其他成员一样，生物合成基因簇中编码的糖基转移酶（ThtS）将N-乙酰葡糖胺添加到ThtA的丝氨酸和半胱氨酸残基上。N-连接的糖基化已被证明在化学和酶促作用下对切割具有抗性，因此ThtS可能是工程研究的一个有价值的起点。对源自ThtA的糖菌素（我们将其命名为嗜热糖菌素）进行了结构表征。嗜热糖菌素的独特之处在于，除了两个嵌套的二硫键外，它还包含一个额外的二硫键，形成了一个C端环。出乎意料的是，ThtA缺乏表示C39肽酶前导肽切割位点的常见双甘氨酸基序。基于AlphaFold3建模，我们推测前导肽和核心肽之间的切割反而发生在GK基序处。这项研究增加了已表征糖菌素的数量，利用AlphaFold3来帮助预测成熟肽产物的结构，并提出了一种类似于为羊毛硫肽建立的通用命名惯例。
