Methods and reagents for site-selective functionalization of peptides and proteins. The methods most generally involve the reaction of a thioester with hydrazine. Reagents include bifunctional reagents of formula:
H
2
N—NH—CH
2
-M-L-FG
and salts thereof where M is a single bond or a chemical group carrying a non-bonding electron pair, such as —C(O)NR′—, where R′ is H, or an alkyl or aryl group; L is an optional linker group as described above; and FG is a functional group having reactivity that is orthongonal to that of the hydrazine group. FG can, among others, be an azide, alkenyl, alkynyl, nitrile (—CN) or triazole group and is preferably an azide group (—N
3
). Methods and reagents can, for example, be combined with intein-mediated protein splicing to link proteins or fragments thereof to various chemical species or to a surface. Surface immobilization of proteins via the methods herein results in immobilized proteins which substantially retain biological activity and is thus useful for the generation of peptide or protein microarrays. Kits for functionalization and/or immobilization of peptides and proteins are provided as well as microarrays of peptides, proteins or both.
肽和蛋白质位点选择性功能化的方法和试剂。这些方法通常涉及将
硫酯与
肼反应。试剂包括双官能团试剂,其
化学式为:
H2N—NH—
CH2-M-L-FG及其盐,其中M是单键或带有非键合电子对的
化学基团,例如—C(O)NR′—,其中R′为H,或烷基或芳基基团;L是上述所述的可选连接基团;FG是具有与
肼基团不同反应性的功能基团。FG可以是偶氮基,烯基,炔基,腈(—CN)或三唑基团等,最好是偶氮基(—N3)。例如,这些方法和试剂可以与内源蛋白质剪切介导的蛋白质剪切相结合,将蛋白质或其片段与各种
化学物种或表面连接起来。通过此处的方法表面固定蛋白质可生成具有
生物活性的固定蛋白质,因此可用于生成肽或蛋白质微阵列。还提供用于肽和蛋白质功能化和/或固定化的试剂盒以及肽,蛋白质或两者的微阵列。