Jul 1, 1976

Synthetic peptides for chymosin and pepsin assays: pH effect and pepsin independent-determination in mixtures

Journal of Dairy Science
R Salesse, J Garnier


Peptide I [H-Phe-Gly-His-Phe(NO2)-Phe-Ala-Phe-OMe] hydrolyzed by chymosin with kcat=.3+/-.3 s-1 and KM=7+/-3 mM (pH 4.7) inhibited competitively peptide II [H-Leu-Ser-Phe(NO2)-Nle-Ala-Leu-OMe] hydrolysis by chymosin with KI=.23 +/- .12 mM at pH 4.7. In reference conditions (.4 mM peptide, .01 M acetate buffer pH 4.7), the specific activities of porcine pepsin and chymosin on peptide I were 470 +/- 70 nM S-1 and .8 nM S-1 per mg of enzyme. This difference in specific activity for peptide I allowed development of a chymosin-independent pepsin assay for mixtures of these enzymes. In addition, peptide II with a specific activity of 2400 +/- 300 nM S-1 and 154 +/- 20 nM S-1 per mg of porcine pepsin and chymosin provides an alternative to measurement of milk clotting for measurement of chymosin- and pepsin-like activities in commercial rennets. Hydrolysis products of peptide II by chymosin exhibited one ionized group of apparent pK of 3.5 +/- .2 and a molar absorption coefficient change of 1000 +/- 100 at pH 4.7 and at 310 nm. From measurements of the kinetic constants, kcat and KM, from pH 2.5 to 7 with peptide II, chymosin activity depends on the protonation of one group of apparent pK 5.3 +/- .2 in the free enzyme. Rennet powder p...Continue Reading

  • References8
  • Citations4


Mentioned in this Paper

Chymosin C
Chymosin Activity
Enzymes, antithrombotic
Structure-Activity Relationship
Pepsin 3
Blood Coagulation Disorders
A 17
Enzymes for Treatment of Wounds and Ulcers
Molar Tooth

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