Prochymosin activation

Procedures for extraction of chymosin from veils were described by Ernstrom and Wong (1974). Crude rennet extract contains active chymosin and an inactive precursor (prochymosin). Addition of acid to the extract facilitates conversion of prochymosin to chymosin and allows the extract to reach maximum activity. Even though activation at lower pH is faster, poor stability of chymosin below pH 5.0 in the pres-... 

Activation of prochymosin involves the splitting of peptides from the N-terminal end of prochymosin with simultaneous reduction in molecular weight from about 36,000 to 31,000. The rate of conversion increases markedly with decreasing pH below 5.0 (Rand and Emstrom 1964). At pH 5.0, NaCl concentrations up to 2M increase the rate of activation. Milk-clotting activity plotted against activation time at pH 5.0 shows the course of activation (Fig. 12.1) to be autocatalytic. If activation is carried out in the presence of preformed chymosin, the S-shape disappears and the initial rate of the activation process increases with increasing concentration of preformed chymosin. Folt-... 

Chymosin (Aspergillus niger var. awamori, Escherichia coli K-12, and Kluyveromyces marxianus, each microorganism containing a calf prochymosin gene), 786, (S3)20 Chymotrypsin, 786, (S3)18 Chymotrypsin Activity, 793 Cinene, 518 1,8-Cineol, 496 Cineole, Percentage of, 818 Cinnamal, 468 Cinnamaldehyde, 468, 611 Cinnamic Acid, 468, 565, 612, (S3)66 Cinnamic Alcohol, 470 Cinnamic Aldehyde, 468 Cinnamon Bark Oil, Ceylon Type, 101, 578... 

Calf prochymosin, calf chymosin (calf prochym., calf chym.). NH2-terminus of the active enzyme is Gly-45. Residues 1-105 (60,61). S-S bridges and residues 357-373 (7). The rest of the sequence from Foltmann and coworkers unpublished results. Residue 290 is aspartic acid in chymosin A and glycine in chymosin B. 

Having considered these difficulties, we have concluded that for the present purpose it is most rational to number the residues from the NH2-terminus of the longest known polypeptide chain of the gastric zymogens (prochymosin) and then continue by counting the longest completely sequenced chain of the active enzymes (porcine pepsin). 

In view of the sequence homologies between pepsinogen and pro-chymosin (Fig.l) it was a little suprising that Al could not inhibit chymosin. This suggested that prochymosin might activate itself by a different mechanism that never generates a 16-residue peptide. 

Activation of Bovine, Canine, and Chicken Pepsinogens and Calf Prochymosin... 

Thus, in the cases of bovine pepsinogen and calf prochymosin where the activation segment sequences are known, peptides were released corresponding to the NH2-terminal parts of the sequence only. For the canine and chicken proteins where no sequence data is available, it appears that inclusion of pepstatin stops the activations before all of the activation segment is released. Consequently, a sequential activation mechanism and not a one-step transformation must be operative with all five of these zymogens. 

By incubation of porcine, bovine, canine, or chicken pepsinogens and calf prochymosin with pepstatin at pH 2.5, the first active protein generated on activation is trapped in an inactive complex. The first activation peptide liberated from porcine pepsinogen has been identified as residues 1-16 whereas that from prochymosin is derived from residues 1-27. This suggests that pepsin and chymosin are not formed by one-step conversions from their zymogens, but by (different) sequential, activation mechanisms. 

V. B. Pedersen and B. Foltmann, University of Copenhagen, Denmark, Comparison of the Activation of Pepsinogen and Prochymosin . 

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