Artichoke (Cynara scolymus L.) flower extract was assayed with the aim of replacing animal rennet in the manufacture of Gouda-type cheeses from bovine milk. Floral extract coagulated milk within a suitable time for use on an industrial scale, while the yield of cheese obtained was equal to that achieved with bovine abomasum. Five proteolytic fractions with milk-clotting activity were isolated in a two-step purification protocol, three belonging to the cardosin group. Cheeses made with C. scolymus proteases must be brined for a longer period (40 h) to prevent overproteolysis and avoid the development of a background flavor. The type of coagulant (bovine or vegetable) had no significant effect on the cheeses' chemical parameters analyzed throughout ripening, and no significant organoleptic differences were detected between those manufactured with C. scolymus or animal rennet. The results indicate that C. scolymus flower extract is suitable for replacing animal rennet in the production of Gouda-type cheeses.
Most of the species belonging to Asclepiadaceae family usually secrete an endogenous milk-like fluid in a network of laticifer cells in which sub-cellular organelles intensively synthesize proteins and secondary metabolites. A new papain-like endopeptidase (asclepain c-II) has been isolated and characterized from the latex extracted from petioles of Asclepias curassavica L. (Asclepiadaceae). Asclepain c-II was the minor proteolytic component in the latex, but showed higher specific activity than asclepain c-I, the main active fraction previously studied. Both enzymes displayed quite distinct biochemical characteristics, confirming that they are different enzymes. Crude extract was purified by cation exchange chromatography (FPLC). Two active fractions, homogeneous by sodium dodecyl sulphate-polyacrylamide gel electrophoresis and mass spectrometry, were isolated. Asclepain c-II displayed a molecular mass of 23,590 Da, a pI higher than 9.3, maximum proteolytic activity at pH 9.4-10.2, and showed poor thermostability. The activity of asclepain c-II is inhibited by cysteine proteases inhibitors like E-64, but not by any other protease inhibitors such as 1,10-phenantroline, phenylmethanesulfonyl fluoride, and pepstatine. The Nterminal sequence (LPSFVDWRQKGVVFPIRNQGQCGSCWTFSA) showed a high similarity with those of other plant cysteine proteinases. When assayed on N-alpha-CBZ-amino acid-p-nitrophenyl esters, the enzyme exhibited higher preference for the glutamine derivative. Determinations of kinetic parameters were performed with N-alpha-CBZ-L-Gln-p-nitrophenyl ester as substrate: K(m)=0.1634 mM, k(cat)=121.48 s(-1), and k(cat)/K(m)=7.4 x 10(5) s(-1)/mM.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.