Butelase 1-Mediated Ligation of Peptides and Proteins
Butelase 1-Mediated Ligation of Peptides and Proteins
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DOI：
10.1007/978-1-4939-9546-2_6
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发表时间：
2019-01-01
期刊：
影响因子：
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通讯作者：
Tam, James P.
中科院分区：
文献类型：
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作者：
Hemu, Xinya;Zhang, Xiaohong;Tam, James P.
在结构上，Butelase 1是天冬酰胺基内蛋白水解酶(AEP)家族的半胱氨酸蛋白酶，但在功能上，它表现出强烈的ASN/Asp特异性(ASX)连接酶活性，几乎不具有蛋白酶活性。Butelase 1在分子内或分子间识别含ASX基序的三肽基序ASX-His-Val，形成ASX-Xaa多肽键(Xaa=任意氨基酸)，分别产生环肽或定点修饰的多肽/蛋白。我们在过去4年的工作已经证明，丁酶1是一种有效的、多功能的多肽和蛋白质修饰工具。在这里，我们描述了使用丁酶1进行高效和定点的多肽和蛋白质连接、N末端标记、硫代酯的制备和树枝状大分子的生物结合的方案。此外，我们还提供了一个使用Butelase 1进行蛋白质环化与遗传密码扩展相结合的例子，以结合非自然的构建块。
Structurally, butelase 1 is a cysteine protease of the asparaginyl endoprotease (AEP) family, but functionally, it displays intense Asn/Asp-specific (Asx) ligase activity and is virtually devoid of protease activity. Butelase 1 recognizes specifically a C-terminal Asx-containing tripeptide motif, Asx-His-Val, to form an Asx-Xaa peptide bond (Xaa = any amino acid), either intramolecularly or intermolecularly, resulting in cyclic peptides or site-specific modified peptides/proteins, respectively. Our work in the past 4 years has validated that butelase 1 is a potent and versatile tool for peptide and protein modification. Here we describe our protocols using butelase 1 for efficient and site-specific peptide and protein ligation, N-terminal labeling, preparation of thioesters, and bioconjugation of dendrimers. Additionally, we provide an example using butelase 1 for protein cyclization in combination with genetic code expansion in order to incorporate unnatural building blocks.