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

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

Subject headings

NATURVETENSKAP  -- Kemi (hsv//swe)

NATURAL SCIENCES  -- Chemical Sciences (hsv//eng)

Keyword

Chemistry

Kemi

Publication and Content Type

ref (subject category)

art (subject category)