Mechanism and structure of thioredoxin reductase from Escherichia coli.
The flavoprotein thioredoxin reductase catalyzes the reduction of the small redox protein thioredoxin by NADPH. Thioredoxin reductase contains a redox active disulfide and is a member of the pyridine nucleotide-disulfide oxidoreductase family of flavoenzymes that includes lipoamide dehydrogenase, glutathione reductase, trypanothione reductase, mercuric reductase, and NADH peroxidase. The structure of thioredoxin reductase has recently been determined from X-ray crystallographic data. In this paper, we attempt to correlate the structure with a considerable body of mechanistic data and to arrive at a mechanism consistent with both. The path of reducing equivalents in catalysis by glutathione reductase and lipoamide dehydrogenase is clear. To envisage the path of reducing equivalents in catalysis by thioredoxin reductase, a conformational change is required in which the NADPH domain rotates relative to the FAD domain. The rotation moves the nascent dithiol from its observed position adjacent to the re surface of the flavin ring system toward the protein surface for dithiol-disulfide interchange with the protein substrate thioredoxin and moves the nicotinamide ring of NADPH adjacent to the flavin ring for efficient hydride transfer. Reverse rotation allows reduction of the redox active disulfide by the reduced flavin. This requires that the enzyme pass through a ternary complex; the kinetic evidence for such a complex is discussed.
Williams
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Department of Veterans Affairs Medical Center, Department of Biological Chemistry, University of Michigan, Ann Arbor 48105, USA.
D002384:Catalysis; D003545:Cysteine / Q000737:chemistry; D008058:Dihydrolipoamide Dehydrogenase / Q000378:metabolism; D004220:Disulfides / Q000737:chemistry; D004926:Escherichia coli / Q000201:enzymology*; D005182:Flavin-Adenine Dinucleotide / Q000378:metabolism; D005420:Flavoproteins / Q000378:metabolism* / Q000648:ultrastructure; D005980:Glutathione Reductase / Q000378:metabolism; D009249:NADP / Q000378:metabolism; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000378:metabolism; D017434:Protein Structure, Tertiary; D013329:Structure-Activity Relationship; D013880:Thioredoxin-Disulfide Reductase / Q000378:metabolism* / Q000648:ultrastructure
D004220:Disulfides; D005420:Flavoproteins; D005182:Flavin-Adenine Dinucleotide; D009249:NADP; D010088:Oxidoreductases; C021551:mercuric reductase; D008058:Dihydrolipoamide Dehydrogenase; D005980:Glutathione Reductase; D013880:Thioredoxin-Disulfide Reductase; D003545:Cysteine