Enzymatic Ligation of Disulfide-Rich Animal Venom Peptides: Using Sortase A to Form Double-Knotted Peptides.
Enzymatic Ligation of Disulfide-Rich Animal Venom Peptides: Using Sortase A to Form Double-Knotted Peptides.
复制标题
富含二硫键的动物毒液肽的酶连接：使用分选酶 A 形成双结肽。
DOI：
10.1007/978-1-0716-1617-8_8
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发表时间：
2021
期刊：
影响因子：
--
通讯作者：
Schroeder,ChristinaI
中科院分区：
文献类型：
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作者：
Tran,Poanna;Schroeder,ChristinaI
分类酶A是一种由革兰氏阳性菌表达的硫醇转肽酶。该酶能够位点特异性地将含有c端识别基序LPXTG的肽连接到含有n端聚甘氨酸序列的肽上，形成天然肽键。在这里，我们描述了分选酶A的制备和应用，以连接两个单独折叠的富含二硫化物的动物毒液肽，以形成具有天然肽连接物的异二聚体双结肽。这种方法足够温和，可以在连接过程中保留肽的结构和二硫连通性。我们使用了一种高效的分选酶a五突变体（SrtA5°），该突变体在15分钟内完成反应，连接肽的产率约为50-80%。
Sortase A is a thiol transpeptidase expressed by Gram-positive bacteria. This enzyme is capable of site-specifically ligating peptides containing the C-terminal recognition motif LPXTG to peptides containing an N-terminal polyglycine sequence, forming a native peptide bond. Here, we describe the preparation and application of sortase A to the ligation of two individually folded disulfide-rich animal venom peptides in order to form a heterodimeric double-knotted peptide with a native peptide linker. This method is mild enough to preserve the structures and disulfide connectivities of the peptides during ligation. We employed a highly efficient sortase A pentamutant (SrtA5°), which brings the reaction to completion within 15 min with a ~50–80% yield of ligated peptide.