A renin inhibitor from rabbit kidney: conversion of a large inactive renin to a smaller active enzyme.

Abstract

Renin in extracts of frozen rabbit kidney exists in two forms: active (molecular weight about 37,000) and inactive (molecular weight about 55,000) renin. The inactive form becomes active after exposure to pH 2.5 at 4°C. If extracts are chromatographed on DEAE cellulose, the inactive renin dissociates into active renin plus a renin inhibitor (molecular weight about 13,000). The inhibitor recombines with active renin if the two are incubated together at 37°C. The inhibitor is destroyed by acid treatment at pH 2.5 at 4°C. We conclude that the activation of inactive renin is due to destruction of the inhibitor by acid. The inactive material may be a renin proenzyme or a storage form of active renin combined with inhibitor. KEY WORDS renin pressor responseproenzyme renin activation molecular weight of renin membrane-bound renin• Extracts of rabbit kidney contain an active and an inactive form of the enzyme renin (1). Inactive rabbit renin has a molecular weight of around 55,000, and active renin has a molecular weight of around 37,000. When a solution of inactive rabbit renin is acidified to pH 2.5, its pressor activity increases and its molecular weight becomes similar to that of active rabbit renin. A slow-acting renin is also present in extracts of pig kidney (2). It is activated by acid and dissociates into active renin plus a "renin-binding protein," when it is chromatographed on DEAE cellulose. The inactive renin from the rabbit kidney may be similar to the slow-acting renin from the pig kidney.The present experiments were carried out in an attempt to characterize the inactive renin from rabbit kidney and to study its activation. MethodsKidney Extracts.-Kidneys were obtained from male or female New Zealand white rabbits. The rabbits were anesthetized with sodium pentobarbital, and their kidneys were excised and stored frozen. The kidney cortex was extracted as previously described (1). Extracts for DEAE cellulose chromatography were made in 0.005M sodium phosphate buffer, pH 7.0, using 3 ml buffer/g kidney cortex.Buffers.-Buffer solutions were boiled and made up to volume in boiled, cooled distilled water. Phosphatesaline buffer contained 0.05M NaHjPO 4 /Na 2 HPO 4 , 0.1M NaCl, and 2.0 g/liter of neomycin sulfate, pH 5.0-7.4. Glycine-HCl buffer contained 0.05M glycine/HCl and 0.1M NaCl, pH 1.5-4.0.From the MRC Blood Pressure Unit, Western Infirmary, Church Street, Glasgow Gil 6NT, Scotland.Received October 7, 1974. Accepted for publication January 13, 1975.Standard Renin.-Standard renin was prepared by a modification of the method of Lever et al. (3). Rabbit kidneys were minced in 0.9% saline and allowed to stand at 4°C for 48 hours. The slurry was centrifuged at 2,000 g for 30 minutes, the supernatant fluid was decanted, the precipitate was washed with half of the original volume of saline, and the washings were added to the supernatant fluid. All subsequent operations were carried out at 4°C. The extract was acidified to pH 2.5 and allowed to stand for 7-16 hours. The pH was adjusted to 7.4, the preci...

Release of active and inactive renin by the porcine kidney.

“…Leckie, 1973;deSenarclens et al, 1977;Skeggs et al, 1967). The studies of Leckie and McConnell (1975) suggest that inactive renin may be coupled to an inhibitor, which is destroyed by acidification. Most studies on experimental animals to date have concentrated on the estimation of tissue renin content, although James and Hall (1974) did measure acidactivated renin in dog plasma.…”

“…This acid-activated or inactive renin* has also been reported to be present in several species in addition to man (Boyd, 1974;Rubin, 1972;Leckie, 1973;deSenarclens et al, 1977;James and Hall, 1974;Skeggs et al, 1967). Whether inactive renin represents a true proenzyme or renin bound to an inhibitor remains unclear (Leckie and McConnell, 1975). It also has been suggested that inactive renin may be the intrarenal storage form of the enzyme (deSenarclens et al, 1977) Acid-activated renin has been measured in systemic and renal venous plasma of man under varying conditions (Derkx et al, 1976;Weinberger et al, 1977).…”

Studies on high molecular weight renin from hog kidney.

“…The reason for this is not immediately apparent, unless the renin-60 is a combination of renin and a bound protein, as reported by Boyd 18 and Leckie and McConnell. 19 If this is the case, it may be that the presence of the protease inhibitors stabilized some other protein which is involved in the conversion of renin-60 to renin-40. If the binding protein in the presence of the protease inhibitors is more labile than renin (as it is at pH 3.3, 18 l9 or possibly even in the cold 12 ' l3 ), relatively more of it would be denatured, thus leaving an insufficient amount to bind all the remaining renin.…”

Characterization of inactive renin ("prorenin") from renin-secreting tumors of nonrenal origin. Similarity to inactive renin from kidney and normal plasma.

“…The unusually high proportion of inactive renin in plasma and tumor extracts from such patients is consistent with ineffective precursor processing by neoplastic tissue, suggesting that if activation of "prorenin" is involved in the normal regulation of active renin levels it more likely Introduction Renin (EC 3.4.99. 19) is a proteolytic enzyme that is found most abundantly in the kidney. Renal renin is secreted into the circulation, where it catalyzes the rate-limiting step in the formation of the vasopressor and aldosterone-stimulating peptide angiotensin II.…”

“…Inactive renin has several characteristics of a biosynthetic precursor of renin (8,12,13). Other evidence has been cited to support the view that it may be a postsynthetic modification ofactive renin (15)(16)(17), especially since, in renal cortical extracts from several species, renin can form a large molecular weight complex with a polypeptide binding substance (18)(19)(20). However, these complexes are enzymatically active and thus differ from the completely inactive substance found in human plasma (8).…”