Engineering the enantioselectivity of glutathione transferase by combined active-site mutations and chemical modifications
2007 (English)In: Biochimica et Biophysica Acta - General Subjects, ISSN 0304-4165, E-ISSN 1872-8006, Vol. 1770, no 9, 1374-1381 p.Article in journal (Refereed) Published
Based on the crystal structure of human glutathione transferase M1-1, cysteine residues were introduced in the substrate-binding site of a Cys-free mutant of the enzyme, which were subsequently alkylated with 1-iodoalkanes. By different combinations of site-specific mutations and chemical modifications of the enzyme the enantioselectivity in the conjugation of glutathione with the epoxide-containing substrates 1-phenylpropylene oxide and styrene-7,8-oxide were enhanced up to 9- and 10-fold. The results also demonstrate that the enantioselectivity can be diminished, or even reversed, by suitable modifications, which can be valuable under some conditions. The redesign of the active-site structure for enhanced or diminished enantioselectivities have divergent requirements for different epoxides, calling for a combinatorial approach involving alternative mutations and chemical modifications to optimize the enantioselectivity for a targeted substrate. This approach outlines a general method of great potential for fine-tuning substrate specificity and tailoring stereoselectivity of recombinant enzymes.
Place, publisher, year, edition, pages
2007. Vol. 1770, no 9, 1374-1381 p.
Enantioselectivity, Epoxide resolution, Glutathione transferase, Protein modification, Rational redesign
IdentifiersURN: urn:nbn:se:uu:diva-11976DOI: 10.1016/j.bbagen.2007.06.002ISI: 000249511200014PubMedID: 17689871OAI: oai:DiVA.org:uu-11976DiVA: diva2:39745