Engineering of a metal coordinating site into human glutathione transferase M1-1 based on immobilized metal ion affinity chromatography of homologous rat enzymes

摘要: Rat glutathione transferase (GST) 3-3 binds to Ni(II)-iminodiacetic acid (IDA)-agarose, whereas other GSTs that are abundant in rat liver do not bind this immobilized metal ion affinity chromatography (IMAC) adsorbent. GST contains two superficially located amino residues, His84 and His85, suitably positioned for coordination Ni(II)-IDA-agarose. This particular structural motif is lacking the IMAC matrix. Creation of an equivalent His-His structure homologous human M1-1 by protein engineering afforded a mutant enzyme displays Ni(II)-IDA-agarose, contrast wild-type M1-1. The results identify distinct site operational suggest approach rational design novel integral sites proteins.