S-carboxymethylcysteine sulfone: instability to acid hydrolysis and unreactivity with N-terminal reagents.

An examination of the properties and reactivity of S-carboxymethylcysteine sulfone indicated that, unlike S-carboxymethylcysteine, the sulfone is not stable under acid hydrolysis conditions and decomposes to yield alanine. Unlike S-carboxymethylcysteine, the sulfone is resistant to N-derivatization by the dansyl reagent or by phenylisothiocyanate. Efforts were made to determine if spontaneous cyclization of the sulfone to the corresponding thiazane (lactam) accounts for lack of reactivity. These included i.r. spectroscopy, natural abundance 13C-n.m.r. spectroscopy and differential scanning calorimetry, but yielded equivocal results concerning the existence of the cyclic form in solution. Solubility behavior of the sulfone after lyophilization from strongly acid solutions was consistent with conversion of the open chain form to the cyclic form on addition of water.

Hydroxyproline-2-epimerase of Pseudomonas: active-site peptides.

Hydroxyproline-2-epimerase was treated with 14C-iodoacetate under conditions that produced almost complete inactivation of the enzyme and concomitant incorporation of almost one molar equivalent of iodoacetate. Both processes were prevented by saturating concentrations of substrate. From reaction mixtures in which both incorporation and inactivation were 85 to 90% complete, two radioactive tryptic peptides were isolated by paper chromatography-electrophoresis. The incorporated radioactivity was divided between the peptides in an approximately 2:1 ratio. Analysis of the isolated peptides suggested that they both contained 9 amino acids and had similar composition; one appeared to be a lysine, the second an arginine peptide. Attempts to sequence each peptide failed, apparently because of the conversion of the S-carboxymethylcysteine to S-carboxymethylcysteine sulfone, indicating that the cysteine residue was N-terminal in each peptide.