MUTANT PYRROLYSYL-TRNA SYNTHETASE, AND METHOD FOR PRODUCTION OF PROTEIN HAVING NON-NATURAL AMINO ACID INTEGRATED THEREIN BY USING THE SAME
申请人:Riken
公开号:EP2192185A1
公开(公告)日:2010-06-02
Method for incorporating a lysine derivative (particularly an N∈-benzyloxycarbonyl-lysine (Z-Lys) derivative) having useful functional group such as heavy atom, selenium, reactive functional group, fluorescent group or crosslinker, which is suitable as a non-natural amino acid, into a desired protein in a site-specific manner.
A mutant pyrrolysyl-tRNA synthetase has substitution of at least one amino acid residue selected from tyrosine residue at position 306, leucine residue at position 309 and cysteine residue at position 348 each constituting a pyrrolysine-binding site in the amino acid sequence for pyrrolysyl-tRNA synthetase of SEQ ID NO:2. The substitution of the amino acid residue is: of tyrosine residue at position 306 by glycine or alanine residue, of leucine residue at position 309 by glycine or alanine residue, and/or of a cysteine residue at position 348 by valine, serine or alanine residue.
将赖氨酸衍生物(特别是 N∈-苄氧羰基-赖氨酸(Z-赖氨酸)衍生物)以特定位点的方式加入到所需蛋白质中的方法,该赖氨酸衍生物具有有用的官能团,如重金属原子、硒、活性官能团、荧光基团或交联剂,适合作为非天然氨基酸。
一种突变体吡咯赖氨酰-tRNA 合成酶至少有一个氨基酸残基被取代,该氨基酸残基选自 SEQ ID NO:2 吡咯赖氨酰-tRNA 合成酶氨基酸序列中分别构成吡咯赖氨酸结合位点的 306 位酪氨酸残基、309 位亮氨酸残基和 348 位半胱氨酸残基。氨基酸残基的取代是:306 位的酪氨酸残基被甘氨酸或丙氨酸残基取代,309 位的亮氨酸残基被甘氨酸或丙氨酸残基取代,和/或 348 位的半胱氨酸残基被缬氨酸、丝氨酸或丙氨酸残基取代。