title: New enzymes for old: redesigning the coenzyme and substrate specificities of glutathione reductase.
abstract: A set of amino acid side chains that confer specificity for the coenzyme NADPH and the substrate glutathione in the flavoprotein disulphide oxidoreductase, glutathione reductase, has been identified. Systematic replacement of these amino acid residues in the coenzyme-binding site switches the specificity of the enzyme from its natural strong preference for NADPH to a marked preference for NADH. The amino acids replaced all lie in a structural motif within the dinucleotide-binding domain of the protein. Since this domain is a feature common to most dehydrogenases (reductases) that use nicotinamide coenzymes, it may be that the coenzyme specificities of all such enzymes can be manipulated in this way. Similarly, amino acid residues involved in the selective recognition of trypanothione by trypanothione reductase, an enzyme related to glutathione reductase and exclusive to trypanosomatids, were identified. Suitable mutation of the corresponding residues in E. coli glutathione reductase switched its substrate specificity towards trypanothione. A better understanding of the substrate specificity of these enzymes could open up a route to the chemotherapy of trypanosomal infections.
authors.affiliation: Department of Biochemistry, University of Cambridge, UK.
authors.forename: R N
authors.initials: RN
authors.lastname: Perham
authors.forename: N S
authors.initials: NS
authors.lastname: Scrutton
authors.forename: A
authors.initials: A
authors.lastname: Berry
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D004926:Escherichia coli / Q000235:genetics; D005980:Glutathione Reductase / Q000235:genetics* / Q000378:metabolism*; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009243:NAD / Q000378:metabolism; D009249:NADP / Q000378:metabolism; D011487:Protein Conformation; D012689:Sequence Homology, Nucleic Acid; D013379:Substrate Specificity
chemical_list: D009243:NAD; D009249:NADP; D005980:Glutathione Reductase