Sortase-mediated ligation (sortagging) is commonly performed using the Staphylococcus aureus sortase A (SaSrtA) that strictly recognizes the N-terminal glycine residue. In this work, a rational design of Streptococcus pyogenes sortase A (SpSrtA) for improved transpeptidase activity toward different N-terminal amino acid residues was conducted. The generated variant SpSrtA M3 (E189H/V206I/E215A) showed up to 6.6-fold (vs SpSrtA wild-type) enhanced catalytic efficiency. Additionally, M3 retains the specificity toward N-terminal alanine, glycine, serine residues, as well as branched (at α-carbon) primary amines as wild-type parent. Furthermore, M3 was applied for head-to-tail backbone cyclization of proteins.

中文翻译：

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设计的化脓性链球菌分选酶A在分选中接受支链胺作为亲核试剂

分选酶介导的连接（分选）通常使用严格识别N末端甘氨酸残基的金黄色葡萄球菌分选酶A（SaSrtA）进行。在这项工作中，进行了化脓性链球菌分选酶A（SpSrtA）的合理设计，以提高针对不同N末端氨基酸残基的转肽酶活性。生成的变体SpSrtA M3（E189H / V206I / E215A）显示高达6.6倍的催化效率（相对于SpSrtA野生型）。另外，M3保留了对N末端丙氨酸，甘氨酸，丝氨酸残基以及支链（在α-碳原子上）伯胺作为野生型亲本的特异性。此外，M3被用于蛋白质从头到尾的主链环化。