Glutathione reductase from human erythrocytes. The sequences of the NADPH domain and of the interface domain.
1. Sequence analysis of the NADPH domain (residues 158--293) and of the interface domain (365--478) was based on 12 CNBr fragments, which were isolated using ion-exchange chromatography and paper methods. Fragments with more than 15 residues were digested further with trypsin and chymotrypsin. The isolated peptides were sequenced by automated solid-phase Edman degradation. All sequenced peptides were ordered and overlapped by computerized comparisons with a complete sequence guessed from the electron density map of the protein. In the case of short CNBr fragments, this alignment was confirmed by the sequence analysis of protein fragments resulting from incomplete CNBr cleavage. 2. In the NADPH domain, residue 197, which is involved in an induced-fit mechanism, was identified as a tyrosine. The structure of the NADPH domain is probably homologous with the NAD domain of lipoamide dehydrogenase and with the FAD domain of several proteins, but not with NADPH domains of known chain-fold in other proteins. 3. The paper completes the sequence analysis of glutathione reductase so that the enzyme is now known in atomic detail. The numbering scheme of the chemically determined sequence will be used henceforth in crystallographic studies also. As inferred from the sequence data each of the two identical chains contains 478 amino acid residues, the composition being Cys10, Asp21, Asn17, Thr31, Ser31, Glu29, Gln11, Pro24, Gly43, Ala42, Val44, Met15, Ile29, Leu34, Tyr13, Phe14, Lys34, His16. Arg17, and Trp3. From these data an Mr of 2 x 51 600 was calculated for the FAD-free apoenzyme and an Mr of 2 x 42 400 for the holoenzyme.
D000595:Amino Acid Sequence; D001665:Binding Sites; D004912:Erythrocytes / Q000201:enzymology*; D005980:Glutathione Reductase / Q000097:blood*; D006801:Humans; D009249:NADP / Q000097:blood*; D010446:Peptide Fragments / Q000032:analysis
D010446:Peptide Fragments; D009249:NADP; D005980:Glutathione Reductase