Identification of a protease inhibitor from rat peritoneal macrophages as poly(ADP-ribose).
Rat peritoneal macrophages contain a chymotrypsin-like protease and its specific inhibitor, both being associated with chromatin of the cells. The inhibitor was separated from the protease by gel filtration through a Sephadex G-75 column, further treated with trypsin, DNase and RNase, and then purified successively on Sephadex G-75, Sephadex G-25, and dihydroxyboryl Bio-Gel P-60 columns. The purified inhibitor had a molecular weight in the range from 2,000 to 3,500 and an absorption maximum at 260 nm at pH 7.0. When the inhibitor was digested by snake venom phosphodiesterase, the inhibitory potency was lost, yielding 5'-AMP and 2'-(5'-phosphoribosyl)-5'-AMP as the digestion products which were identified by high pressure liquid chromatography. The inhibitory potency was neutralized specifically by anti-poly(ADP-ribose) antiserum. The profile of inhibition by the isolated inhibitor was nearly identical with that produced by authentic poly(ADP-ribose). It was therefore concluded that the inhibitor isolated was identical with poly(ADP-ribose), whose chain length ranged from 4 to 7 ADP-ribosyl units. This is the first demonstration that a intracellular protease inhibitor can be endogenous poly(ADP-ribose).
D000818:Animals; D001202:Ascitic Fluid / Q000166:cytology; D002843:Chromatin / Q000032:analysis; D007106:Immune Sera; D008264:Macrophages / Q000378:metabolism*; D008297:Male; D009702:Nucleoside Diphosphate Sugars / Q000302:isolation & purification*; D010727:Phosphoric Diester Hydrolases / Q000378:metabolism; D011064:Poly Adenosine Diphosphate Ribose / Q000276:immunology / Q000302:isolation & purification* / Q000494:pharmacology; D011480:Protease Inhibitors / Q000302:isolation & purification*; D051381:Rats; D012910:Snake Venoms; D013056:Spectrophotometry, Ultraviolet
D002843:Chromatin; D007106:Immune Sera; D009702:Nucleoside Diphosphate Sugars; D011480:Protease Inhibitors; D012910:Snake Venoms; D011064:Poly Adenosine Diphosphate Ribose; D010727:Phosphoric Diester Hydrolases