Prorenins activation by an enzyme from rat plasma (PreR-Co) (original) (raw)

Rat Renal and Plasma Prorenin Are Activated In Vitro by Different Mechanisms

Hypertension, 1999

The aim of the present study was to purify and identify a plasma protein fraction (PreR-Co) involved in renal prorenin activation and to explore its capacity to process plasma prorenin. PreR-Co was obtained from plasma as a single electrophoretic band by (NH 4) 2 SO 4 precipitation, Sephacryl S-200 HR gel filtration, anti-rat albumin immunoaffinity, and ion-exchange chromatography. The amidase, esterase, and kallikrein activities of PreR-Co were studied, as was its N-terminal amino acid sequence. Rat kidney extract or plasma (normal or previously treated with acid to pH 2.8) were incubated with PreR-Co for 15 minutes at 37°C. Renin concentration was measured by incubation with homologous angiotensinogen. The same protocol was repeated with samples activated by trypsin. The N-terminal amino acid sequence was IIGGSMDAKGSFP, which had a homology of 90% with the ␤-chain of haptoglobin, 69% with serine-proteases, and 65% with kallikreins. The renin concentration in rat kidney extract was 34Ϯ4 ng of angiotensin I (Ang I) ⅐ mg of tissue Ϫ1 ⅐ h Ϫ1. After PreR-Co or trypsin treatments, renin concentrations were 211Ϯ7 and 110Ϯ11 ng of Ang I ⅐ mg of tissue Ϫ1 ⅐ h Ϫ1 , respectively. The plasma renin concentration in normal plasma was 67.6Ϯ13.3 ng of Ang I ⅐ mL Ϫ1 ⅐ h Ϫ1 , and no significant difference was observed after PreR-Co treatment. However, a significant increase (202.8Ϯ7.8 ng of Ang I ⅐ mL Ϫ1 ⅐ h Ϫ1 ; PϽ0.01) was found after trypsin treatment. The isolated PreR-Co acts on renal prorenin but not on plasma prorenin. These results suggest that active renin is processed in the kidney by a circulating enzyme that may have a role in the regulation of circulating renin. (Hypertension. 1999;34:520-524.

Measurement and Identification of Prorenin and Renin in Ovarian Follicular Fluid from Cattle and Pig

Clinical and Experimental Pharmacology and Physiology, 1992

1. In previous studies we have demonstrated and solved several methodological problems in relation to the measurement of prorenin by trypsin activation in rat, bovine, hog and horse plasma. 2. The aim of the present study was to develop a method for the measurement of prorenin in bovine and porcine ovarian follicular fluid. 3. Trypsin activation of follicular fluid generated angiotensin I immunoreactive material (A1 IM) in both species. 4. The A1 IM interfered with the renin assay, but could be completely removed by a cation exchange resin in a batch-wise technique. 5. The enzymatic activity of trypsin-activated prorenin and pre-existing active renin was completely inhibited by a specific inhibitor of renin. 6. The reactions were optimized and an accurate measurement of prorenin in ovarian follicular fluid was developed. 7. The existence of prorenin and renin in bovine ovarian follicular fluid was established. Prorenin and renin in porcine ovarian follicular fluid was demonstrated for the first time. 8. The ratio between ovarian follicular fluid and plasma was 43 for prorenin and 19 for active renin in cattle. The same ratios in pigs were 1.3 and 0.4, respectively. These findings indicate a species difference with respect to the amount of prorenin or active renin present in ovarian follicular fluid.

Proteolytic processing of human prorenin in renal and non-renal tissues

Kidney International, 1994

Proteolytic processing of human prorenin in renal and non-renal tissues. Previous studies have demonstrated that the mouse proprotein convertase PCi (mPC1) accurately cleaves human prorenin to generate active renin and that this processing event appears to require co-packaging in secretory granules. In the current study, we have tested human PCi (hPC1; also called PC3) for its ability to activate human prorenin. Our results suggest that while hPC1 is capable of carrying out the specific cleavage of human prorenin, it does so at a reduced efficiency as compared to mPC1. This difference is due to sequences in the carboxy-terminus of PCi as demonstrated by the activity of hybrid hPC1/mPC1 molecules. These studies demonstrate that PCi cleavage of prorenin can occur in humans and identify a functionally important region in the hPCi protein for this interaction. Moreover, the localization of PCi in human tissues suggests that it may participate in the generation of active renin in the adrenal medulla and possibly in certain adrenal tumors.

Improved immunoradiometric assay for plasma renin

Clinical chemistry, 1999

Our renin IRMA overestimated renin in plasmas with high prorenin-to-renin ratios. We suspected that the overestimation of renin was caused less by cross-reactivity of the renin-specific antibody with prorenin than by a conformational change of prorenin into an enzymatically active form during the assay. Because the inactive form of prorenin converts slowly into an active form at low temperature, we raised the assay temperature from 22 degrees C to 37 degrees C, simultaneously shortening the incubation time from 24 to 6 h. The former IRMA was performed in <1 working day with these modifications. The comeasurement of prorenin as renin was eliminated. Reagents were stable at 37 degrees C, and the new and old IRMAs were comparable in terms of precision and accuracy. The functional lower limit of the assay (4 mU/L) was below the lower reference limit (9 mU/L). The modified IRMA agreed closely with the activities measured with an enzyme-kinetic assay. Results were not influenced by the...

Species specificity of prorenin binding to the (pro)renin receptor in vitro

Frontiers in Bioscience, 2010

Introduction 3. Materials and Methods 3.1. Preparation of human and rat prorenin 3.2. Expression of human and rat (pro)renin receptor on the membrane of COS-7 cells 3.3. Bindings of prorenin molecules to the (pro)renin receptor expressed on the membrane of COS-7 cells 3.4. In vitro expression of recombinant human and rat (pro)renin receptor 3.5. Binding assay of prorenin molecules to the (pro)renin receptor using BIAcore assay system 3.6. Determination of the molecular activities of receptor-bound prorenin molecules 4. Results 4.1. Binding of prorenin to (pro)renin receptor expressed on the membrane of COS-7 cells 4.2. Real-time bindings of prorenin molecules to the immobilized (pro)renin receptors in BIAcore 4.3. Molecular activities of the receptor-bound prorenin 5. Discussion 6. Acknowledgements 7. References

Pure human inactive renin. Evidence that native inactive renin is prorenin

The Journal of biological chemistry, 1989

To clarify contradicting observations on the identity of inactive renin and prorenin, inactive renin was completely purified from native human chorion laeve and the culture medium of human chorion cells. A 720,000-fold purification with 14% recovery was achieved from chorion laeve in 6 steps, including immunoaffinity chromatography on a monoclonal antibody to human renin coupled to Protein A-Sepharose CL-4B. A 3,100-fold purification with 40% recovery was achieved from chorion culture medium in 4 steps, including immunoaffinity chromatography. Inactive renin purified from the two different sources migrated as a single protein band with the same molecular weight of 47,000 on sodium dodecyl sulfate-polyacrylamide gel electrophoresis and consisted of multiple components that could be resolved by isoelectric focusing. Both had the same pI values which shifted downward upon activation by trypsin; however, relative peak heights were different between the two preparations. The purified ina...

Prorenin as a reproductive hormone. New form of the renin system

The American Journal of Medicine, 1986

Prorenin, the biosynthetic precursor of renin, is synthesized by the kidneys. Herein is reviewed recent evidence that the ovaries also secrete prorenin. It was found that prorenin is present in mature human ovarian follicular fluid in extremely high concentrations and that plasma prorenin levels increase transiently in blood during the menstrual cycle at the time of ovulation. No change in plasma active renin levels occurs at this time. Plasma prorenin level also increases lo-fold in pregnant women very soon after conception. The ovaries are apparently the source of this rise, since plasma prorenin levels did not increase in a pregnant woman with ovarian failure who received a donor egg. All of these changes in plasma prorenin levels appear to be caused by gonadotropic hormones. These results suggest a role for ovarian prorenin in human reproductive function. They may have relevance to studies of female infertility, birth control, and toxemia of pregnancy. They also suggest that a renin system exists that is regulated by changes in prorenin. Renin is classically considered to be an enzyme that is synthesized by the kidneys and secreted into the circulation where it cleaves angiotensin I from angiotensinogen. Angiotensin I has no known physiologic effect but is converted to the octapeptide angiotensin II by converting enzyme, a dipeptidase that is ubiquitous but that is present in very high concentrations in the lung. Angiotensin II is the active hormone of the reninangiotensin system, causing primarily arteriolar vasoconstriction and increased adrenal aldosterone biosynthesis [I].

Biochemical properties of renin and prorenin binding to the (pro)renin receptor

Hypertension Research, 2009

The discovery of (pro)renin receptor, (P)RR, has made the renin-angiotensin system (RAS) more multifaceted. Interaction of renin and prorenin with this receptor has set a new perspective about the physiological functions, activation mechanism and pathophysiological roles of renin/prorenin. Uses of peptides mimicking the structure of the ligands have been very effective for determining structure-function relationship between the ligands and receptor. The probable pivotal role of decoy peptide region (R 10P IFLKRMPSI 19P) of prorenin prosegment was suggested for higher binding affinity of prorenin to (P)RR than that of mature renin. Recently, 'hinge' region peptide (S 149 QGVLKEDVF 158) in renin/prorenin molecule has been reported. Both renin and prorenin can interact with (P)RR through the 'hinge' region. Furthermore, it has been proposed that prorenin has multiple binding sites whereas renin has a single binding site for (P)RR. To comprehend the activation mechanism of renin and prorenin after receptor binding, it is very important to understand their interaction with the receptor. Several kinds of peptides designed from the regions of the tertiary structure of renin and predicted model of prorenin facilitated the study of the in vitro binding mechanisms for renin and prorenin to (P)RR. Here, a series of recent in vitro studies was reviewed to discuss a possible binding mechanism of renin/prorenin to the (P)RR.