Partial Characterization of Hog Renin Purified by Affinity Chromatography

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Abstract
A method was developed to purify [hog] renin [EC 3.4.99.19] on a large scale by chromatography using Pepstatin, a potent inhibitor of renin, as a ligand. Pepstatin was covalently coupled to Sepharose via 6 different spacer ''arms''. The Sepharose-hexamethylenediamino-Pepstatin was the better derivative for renin purification even at a concentration as low as 160 nmol of Pepstatin/ml of moist gel. Renin was extracted from 100 kg of hog kidneys and semi-purified by ammonium sulfate precipitations and chromatography on DEAE-cellulose. The active fraction (48.5 g of proteins) was applied on a 500-ml column. Renin was eluted in the starting buffer containing 6 M urea. Renin was purified 120-fold by the chromatography step with a 79% recovery. Physico-chemical characterization of highly purified renin was performed. Isoelectrofocusing on a pH gradient from 3 to 6 showed a major peak with an isoelectric point (pI) of 4.95 and a minor peak (pI = 4.70). Polyacrylamide gel electrophoresis, pH 7.8, at different gel concentrations, showed a single peak of renin activity which was in the major protein band. Molecular size estimated on agarose-acrylamide gel filtration was 40,000. The physical parameters were similar before and after purifiation.