The Isolation and Primary Structure of a Peptide Containing the Oxidation-Reduction Active Cystine of Escherichia coli Lipoamide Dehydrogenase

An 11-residue peptic peptide from the catalytic center of Escherichia coli lipoamide dehydrogenase has been isolated and sequenced. This peptide contains the oxidation-reduction active disulfide of the enzyme which functions in concert with the flavin during catalysis. The peptide was isolated from a digest of a derivative in which the free sulfhydryls of the enzyme were substituted with the colored maleimide N-(4-dimethylamino-3,5-dinitrophenyl)maleimide. The sequence of the peptide was determined by a subtractive Edman degradation to be: [see PDF for sequence] This work suggests that the oxidation-reduction active disulfide of E. coli lipoamide dehydrogenase is contained in a hydrophobic pocket at the catalytic center of the enzyme, consistent with the binding of the nonpolar substrate dihydrolipoamide. Evidence is also presented that the two subunits of the enzyme are identical.