Amino acids yields during proteolysis catalyzed by carboxypeptidase A are strongly dependent on substrate pre-hydrolysis

The hydrolysis of polypeptides (45 °C, pH 7.0), catalyzed by bovine pancreas carboxypeptidase A (CPA) immobilized on 6%-agarose beads (weight basis), activated with linear aliphatic aldehyde groups (glyoxyl-agarose) is studied here. Reaction substrates (polypeptides) were the product of previous sequential hydrolyses of cheese whey proteins by trypsin and/or chymotrypsin. Yields of individual amino acids were correlated with respect to the overall rate of hydrolysis, thus allowing the prediction of the rates of production of each amino acid during the reaction course. Two kinetic models were compared, resulting respectively from a classic Briggs–Haldane formulation and from the relaxation of the pseudo-steady-state assumption. This approach grants lumped-parameter models, since it considers the number of hydrolysable peptide bounds as a pseudo-component, and in the presence of mass transport effects within the biocatalyst pores. Yet, our methodology was able to provide a detailed composition of a tailor-made pool of amino acids and peptides, allowing the optimization of the enzymatic reactor operation. It was observed that different conditions of substrate pre-hydrolysis caused significant changes on these yields. CPA hydrolyzed up to 11 amino acids, but its activity was higher for Phe, followed by Leu, Tyr and Trp.