St-PepA, a aminopeptidase with high specificity for acidic residues Free

Abstract

The proteolytic system of lactic acid bacteria has been extensively studied over the past 10 years and peptidases from lactococci are now well known. The situation is, however, different for from which only a few peptidases have been purified and characterized. The present work was conducted to characterize an aminopeptidase of CNRZ 302, called St-PepA, with high specificity for acidic amino acids. St-PepA was purified by a three-step procedure from a spheroblast extract of CNRZ 302. Its molecular mass was estimated to be 360 kDa by gel filtration and 45 kDa by SDS-PAGE, indicating that it had an octameric structure. Its activity against aspartate-p-nitroanilide was maximal at pH 8.5 and 62 C and highly enhanced by Zn, Co and . St-PepA was inhibited by metal-chelating reagents and, to a lesser extent, by agents modifying sulfhydryl groups. It showed an activity towards p-nitroanilide derivatives, di-and tripeptides, and larger peptides such as fragment 43-58 of a-casein. It had a high substrate specificity towards N-terminal acidic amino acid residues but it could also release serine and malic acid, the -hydroxy acid homologue of aspartic acid. Kinetic studies revealed that the affinity of St-PepA was more than 18-fold higher for aspartic acid--nitroanilide ( = 0.42 mM) than for glutamic acid--nitroanilide ( = 7.65 mM) with a similar for both substrates [about 40 mol min (mg enzyme)]. St-PepA is heat stable, with a maximal loss of activity of 15% after incubation for 120 min at 50 C and 60C. It is likely to be involved in the nitrogen metabolism of and in the development of the organoleptic characteristics of cheese and yoghurt.

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1995-09-01
2024-03-28
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