Modification of N-Terminal α-Amino Groups of Peptides and Proteins Using Ketenes
Modification of N-Terminal α-Amino Groups of Peptides and Proteins Using Ketenes
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DOI：
10.1021/ja208009r
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发表时间：
2012-02-08
影响因子：
15
通讯作者：
Che, Chi-Ming
中科院分区：
文献类型：
--
作者：
Chan, Anna On-Yee;Ho, Chi-Ming;Che, Chi-Ming
建立了一种用分离的烯酮对蛋白质和多肽进行高选择性N-末端修饰的方法。在室温、pH 6.3的条件下，用炔基功能化的烯酮(1)修饰未受保护的多肽文库XSKFR(X变化超过20个天然氨基酸)，20个多肽中的13个具有良好的N端选择性(修饰的α-氨基/修饰的β-氨基=>；99：1)，其余7个多肽中的6个具有中等到高的N-末端选择性(4：1至48：1)。用炔基功能化的N-羟基丁二酰亚胺(NHS)酯(2)代替1，对多肽XSKFR进行修饰，得到了20个多肽中的5个内部赖氨酸修饰的多肽和13个中低N端选择性(5：1~1：4)的多肽。包括胰岛素、溶菌酶、RNaseA和治疗性蛋白质BCArg在内的蛋白质在室温下被烯酮1选择性地N-末端修饰，而在NHS酯2的修饰中形成了大量或主要的二、三或四修饰蛋白质。1-修饰的蛋白质通过点击化学被丹磺酰叠氮化合物进一步官能化，而不需要事先处理。
A method of highly selective N-terminal modification of proteins as well as peptides by an isolated ketene was developed. Modification of a library of unprotected peptides XSKFR (X varies over 20 natural amino acids) by an alkyne-functionalized ketene (1) at room temperature at pH 6.3 resulted in excellent N-terminal selectivity (modified alpha-amino group/modified epsilon-amino group = >99:1) for 13 out of the 20 peptides and moderate-to-high N-terminal selectivity (4:1 to 48:1) for 6 of the 7 remaining peptides. Using an alkyne-functionalized N-hydroxysuccinimide (NHS) ester (2) instead of 1, the modification of peptides XSKFR gave internal lysine-modified peptides for 5 out of the 20 peptides and moderate-to-low N-terminal selectivity (5:1 to 1:4) for 13 out of the 20 peptides. Proteins including insulin, lysozyme, RNaseA, and a therapeutic protein BCArg were selectively N-terminally modified at room temperature using ketene 1, in contrast to the formation of significant or major amounts of di-, tri-, or tetra-modified proteins in the modification by NHS ester 2. The 1-modified proteins were further functionalized by a dansyl azide compound through click chemistry without the need for prior treatment.