Evidence for reactivity of serine-74 with trans-4-(N,N-dimethylamino)cinnamaldehyde during oxidation by the cytoplasmic aldehyde dehydrogenase from sheep liver

Publisher Website
Google Scholar
Abstract
A nucleophilic group in the active site of aldehyde dehydrogenase, which covalently binds the aldehyde moiety during the enzyme-catalyzed oxidation of aldehydes to acids, was acylated with the chromophoric aldehyde trans-4-(N,N-dimethylamino)cinnamaldehyde (DACA). Acyl-enzyme trapped by precipitation with perchloric acid was digested with trypsin, and the peptide associated with the chromophoric group was isolated and shown to be Gln-Ala-Phe-Gln-Ile-Gly-Ser-Pro-Trp-Arg. After redigestion with thermolysin, the chromophore was associated with the C-terminal hexaresidue part. If the chromophore is attached to this peptide, serine would be expected to bind the aldehyde and lead to the required acylated derivative. Differential labeling experiments were performed in which all free thiol groups on the acylated enzyme were blocked by carboxymethylation. The acyl chromophore was then removed by controlled hydrolysis and the protein reacted with [14C]iodoacetamide. No 14C-labeled tryptic peptides were isolated, suggesting that the sulfur of a cysteine cannot be the acylated residue in the precipitated acyl-enzyme.