Tight-binding inhibition of angiogenin and ribonuclease A by placental ribonuclease inhibitor.
The dissociation rate constant of the angiogenin-placental ribonuclease inhibitor complex was determined by measuring the release of free angiogenin from the complex in the presence of scavenger for free placental ribonuclease inhibitor (PRI). In 0.1 M NaCl, pH 6, 25 degrees C, this value is 1.3 X 10(-7) s-1 (t1/2 congruent to 60 days). The Ki value for the binding of PRI to angiogenin, calculated from the association and dissociation rate constants, is 7.1 X 10(-16) M. The corresponding values for the interaction of RNase A with PRI, determined by similar means, are both considerably higher: the dissociation rate constant is 1.5 X 10(-5) s-1 (t1/2 = 13 h), and the Ki value is 4.4 X 10(-14) M. Thus, PRI binds about 60 times more tightly to angiogenin than to RNase A. The effect of increasing sodium chloride concentration on the binding of PRI to RNase A was explored by Henderson plots. The Ki value increases to 39 pM in 0.5 M NaCl and to 950 pM in 1 M NaCl, suggesting the importance of ionic interactions. The mode of inhibition of RNase A by PRI was determined by examining the effect of a competitive inhibitor of RNase A, cytidine 2'-phosphate, on the association rate of PRI with RNase A. Increasing concentrations of cytidine 2'-phosphate decrease the association rate in a manner consistent with a competitive mode of inhibition.
Lee
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Center for Biochemical and Biophysical Sciences and Medicine, Harvard Medical School, Boston, Massachusetts 02115.
Shapiro
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Vallee
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D043925:Angiogenesis Inducing Agents / Q000037:antagonists & inhibitors* / Q000378:metabolism; D001667:Binding, Competitive; D006131:Growth Inhibitors; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D010926:Placental Hormones / Q000378:metabolism*; D011506:Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D012259:Ribonuclease, Pancreatic / Q000037:antagonists & inhibitors* / Q000378:metabolism
D043925:Angiogenesis Inducing Agents; D006131:Growth Inhibitors; D010926:Placental Hormones; D011506:Proteins; C022052:placental ribonuclease inhibitor; C046366:angiogenin; D012259:Ribonuclease, Pancreatic