CHARACTERIZATION OF CARBOXYPEPTIDASE ACTI VITY OF PENICILLIUM-CAMEMBERTI

In the present study characteristics of carboxypeptidase activity of P enicillium camemberti, are given, and production levels of free enzyme s in culture medium or still bound to mycelium are determined. Strains were grown on Czapeck trypticase medium with initial pH 4.0 or 6.5. T he culture medium after mycelium separation was the exocellular prepar ation. Mycelium was washed with water and then ground in a buffer; it contained 1% Triton X100. This suspension was defined as the raw endoc ellular preparation. After 40,000 g centrifugation for 1 h, the supern atant represented the endocellular soluble extract. Activities were me asured at pH 4.0 on carbobenzoxy-glutamyl tyrosine (Z-glu-tyr) and at pH 6.0 or 7.5 on carbobenzoxy-glycyl-valine (Z-gly-val). Exo- or endoc ellular preparations were applied to a Sephadex G-150 column after con centration by ultrafiltration. Results show that carboxypeptidase acti vity arises from at least 3 fractions. Exocellular activity could be i dentified as a carboxypeptidase acid, its optimal pH was 3.5, its maxi mum stability between pH 4.0 and 5.0 and its molecular weight was abou t 120,000. It was particularly active on peptides with aromatic acid i n C-terminal position; the exocellular activity corresponded to that o f an enzyme previously described. The mycelium bound activity, solubil ized by Triton X100, consisted in 2 fractions. The first one, precipit able by acetone, showed no activity on Z-glu-tyr and Z-glu-phe by-prod ucts and was active on Z-gly-val between pH 5.0 and 9.0. The second on e, unprecipitable (or denatured) by acetone, preferably hydrolysed the same substrates as exocellular enzyme, but its optimum pH was 5.5-6.0 . The production study performed on 16 strains selected from a collect ion of hundred strains showed a limited amplitude (1 to 3) of variatio n. Penicillium camemberti carboxypeptidase activity comprises one exoc ellular acid carboxypeptidase and two neutral enzymes, endocellular or still bound to mycelium. Their characteristics could be taken into ac count in cheese ripening control.