The function of the milk-clotting enzymes bovine and camel chymosin studied by a fluorescence resonance energy transfer assay

Journal of Dairy Science
Jesper Langholm JensenJohannes M van den Brink

Abstract

Enzymatic coagulation of bovine milk can be divided in 2 steps: an enzymatic step, in which the Phe105-Met106 bond of the milk protein bovine κ-casein is cleaved, and an aggregation step. The aspartic peptidases bovine and camel chymosin (EC 3.4.23.4) are typically used to catalyze the enzymatic step. The most commonly used method to study chymosin activity is the relative milk-clotting activity test that measures the end point of the enzymatic and aggregation step. This method showed that camel chymosin has a 2-fold higher milk-clotting activity toward bovine milk than bovine chymosin. To enable a study of the enzymatic step independent of the aggregation step, a fluorescence resonance energy transfer assay has been developed using a peptide substrate derived from the 98-108 sequence of bovine κ-casein. This assay and Michaelis-Menten kinetics were employed to determine the enzymatic activity of camel and bovine chymosin under milk clotting-like conditions (pH 6.65, ionic strength 80 mM). The results obtained show that the catalytic efficiency of camel chymosin is 3-fold higher than bovine chymosin. The substrate affinity and catalytic activity of bovine and camel chymosin increase at lower pH (6.00 and 5.50). The glycosylatio...Continue Reading

References

Nov 1, 1969·The Biochemical Journal·B Foltmann
Sep 12, 2002·The Journal of Dairy Research·Maurice G HayesAlan L Kelly
Feb 21, 2006·Biochemical and Biophysical Research Communications·Stefan R KappelerEric Johansen
May 2, 2013·Acta Crystallographica. Section D, Biological Crystallography·Jesper Langholm JensenSine Larsen

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