Pure Renin. Isolation from hog kidney and characterization.
The pressor enzyme renin (EC 3.4.99.19) was isolated in a pure and stable form from hog kidney by affinity chromatography on a pepstatin/agarose gel followed by three additional steps of conventional chromatography. Destruction of the enzyme by proteolysis during isolation was prevented by chemically eliminating proteases in extracts. The pure preparation was used for the characterization of this enzyme. Renin was found to be a glycoprotein containing glucosamine and possessing binding affinity to concanavalin A. Contrary to previous reports, pure renin is stable at neutral pH either at 4 or -20 degrees for 3 to 8 weeks. It has a molecular weight of 36,400 as determined by equilibrium ultracentrifugation, an isoelectric point of 5.2 and E1%1cm (280 nm) of 9.1. In contrast to crude preparations, the enzyme activity has a broad pH optimum between pH 5.5 and 7.0 for both hog angiotensinogen and the synthetic octapeptide substrate benzyloxycarbonyl-Pro-Phe-His-Leu-Leu-Val-Tyr-Ser-beta-naphthylamide. The rate of formation of angiotensin I from hog angiotensinogen at pH 6.0 and 37 degrees was 267 microng/h/microng of renin, or 2000 Goldblatt units/mg of renin. For the synthetic fluorogenic octapeptide substrate benzyloxycarbonyl-Pro-Phe-His-Leu-Leu-Val-Tyr-Ser-beta-naphthylamide, a Km of 33 micronM and a Vmax of 0.94 micronmol/h/mg of enzyme were obtained at pH 6.5 and 37 degrees.
D000808:Angiotensinogen / Q000378:metabolism; D000818:Animals; D055598:Chemical Phenomena; D002621:Chemistry; D003208:Concanavalin A / Q000378:metabolism; D005944:Glucosamine / Q000032:analysis; D006023:Glycoproteins / Q000032:analysis; D006863:Hydrogen-Ion Concentration; D007668:Kidney / Q000201:enzymology*; D007700:Kinetics; D008970:Molecular Weight; D009842:Oligopeptides / Q000378:metabolism; D011485:Protein Binding; D012083:Renin / Q000302:isolation & purification* / Q000378:metabolism; D013552:Swine
D006023:Glycoproteins; D009842:Oligopeptides; D000808:Angiotensinogen; D003208:Concanavalin A; D012083:Renin; D005944:Glucosamine