TY - JOUR
T1 - The Effect of Limited Proteolysis of Chicken Ovoinhibitor by Bovine Chymotrypsin on the Inhibitory Activities against Trypsin, Chymotrypsin and Elastase
AU - Feinstein, Gad
AU - Gertler, Arieh
PY - 1972/11
Y1 - 1972/11
N2 - Incubation of chicken ovoinhibitor with catalytic amounts of bovine α‐chymotrypsin at low pH caused partial losses in the inhibitory activities against bovine chymotrypsin and porcine elastase, but not against bovine trypsin. Similar incubation with trypsin or elastase, even followed by incubation with carboxypeptidase A or B, had no effect on any inhibitory activity. The losses of the inhibitory activities were pH dependent and had different pH profiles for the effect on the inhibitory activities against chymotrypsin and elastase. The maximal losses were observed around pH 3, while incubation at pH 5 still resulted in 40% loss of chymotrypsin‐inhibiting activity with no effect on elastase‐inhibiting activity. The inhibitory activity against chymotrypsin was also lost faster, thus indicating that the inhibitory sites against these enzymes are distinct and differ from each other. The N‐ and C‐terminal amino acid analyses of ovoinhibitor treated with chymotrypsin at pH 3 and 5, as compared to the native ovoinhibitor, indicate a cleavage of Phe‐Ile bond that could be related to the partial loss of chymotrypsin‐inhibiting activity. The cleavage of Tyr‐X bond was observed mainly after treatment at pH 3 and therefore was related to the loss of elastase‐inhibiting activity. The N‐terminal of the native ovoinhibitor was found to be isoleucine. The inhibitory activities of chicken ovoinhibitor that was isolated by affinity chromatography on chymotrypsin‐Sepharose were compared to those of ovoinhibitor that was isolated by salt fractionation. The trypsin‐inhibitory activities of both preparations were equal, but the chymotrypsin and elastase‐inhibitory activities were lower than those of salt‐fractionated ovoinhibitor. These results, together with the N‐ and C‐terminal amino acid analyses, indicate that during the affinity chromatography on insoluble chymotrypsin, there occurred a limited proteolysis of the ovoinhibitor.
AB - Incubation of chicken ovoinhibitor with catalytic amounts of bovine α‐chymotrypsin at low pH caused partial losses in the inhibitory activities against bovine chymotrypsin and porcine elastase, but not against bovine trypsin. Similar incubation with trypsin or elastase, even followed by incubation with carboxypeptidase A or B, had no effect on any inhibitory activity. The losses of the inhibitory activities were pH dependent and had different pH profiles for the effect on the inhibitory activities against chymotrypsin and elastase. The maximal losses were observed around pH 3, while incubation at pH 5 still resulted in 40% loss of chymotrypsin‐inhibiting activity with no effect on elastase‐inhibiting activity. The inhibitory activity against chymotrypsin was also lost faster, thus indicating that the inhibitory sites against these enzymes are distinct and differ from each other. The N‐ and C‐terminal amino acid analyses of ovoinhibitor treated with chymotrypsin at pH 3 and 5, as compared to the native ovoinhibitor, indicate a cleavage of Phe‐Ile bond that could be related to the partial loss of chymotrypsin‐inhibiting activity. The cleavage of Tyr‐X bond was observed mainly after treatment at pH 3 and therefore was related to the loss of elastase‐inhibiting activity. The N‐terminal of the native ovoinhibitor was found to be isoleucine. The inhibitory activities of chicken ovoinhibitor that was isolated by affinity chromatography on chymotrypsin‐Sepharose were compared to those of ovoinhibitor that was isolated by salt fractionation. The trypsin‐inhibitory activities of both preparations were equal, but the chymotrypsin and elastase‐inhibitory activities were lower than those of salt‐fractionated ovoinhibitor. These results, together with the N‐ and C‐terminal amino acid analyses, indicate that during the affinity chromatography on insoluble chymotrypsin, there occurred a limited proteolysis of the ovoinhibitor.
UR - http://www.scopus.com/inward/record.url?scp=0015519016&partnerID=8YFLogxK
U2 - 10.1111/j.1432-1033.1972.tb02495.x
DO - 10.1111/j.1432-1033.1972.tb02495.x
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C2 - 4674348
AN - SCOPUS:0015519016
SN - 0014-2956
VL - 31
SP - 25
EP - 31
JO - European Journal of Biochemistry
JF - European Journal of Biochemistry
IS - 1
ER -