Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.21.4 (trypsin)
42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Cleavage of prorenin's prosegment causes irreversible formation of renin. In contrast, renin activity is reversibly exposed when prorenin is acidified to pH 3.3. Nonetheless, acidification of plasma results in irreversible activation of prorenin, because endogenous proteases cleave the prosegment of acid-activated prorenin. Chilling of plasma results in irreversible cryoactivation of prorenin. In this study we investigated whether cryoactivation of purified prorenin is reversible. The intrinsic renin activity of recombinant human prorenin was measured by an enzyme kinetic assay using partially purified human angiotensinogen as substrate. Results are expressed as a percent (mean +/- S.E.) of the maximal activity exposed after limited proteolysis by trypsin. The intrinsic renin activity of two pools (0.3 and 0.06 Goldblatt units/ml) was 1.5% +/- 0.3 and 1.2% +/- 0.6 at 37 degrees C. Activity increased to 19% +/- 0.3 and 26% +/- 0.5 after incubation at 0 degrees C and to 5.4% +/- 0.5 and 2.1% +/- 1.2 at room temperature. Cryoactivation did not occur in buffers containing more than 1 M NaCl. It took 8 min at 37 degrees C or 180 min at room temperature for cryoactivated prorenin to lose half of its intrinsic renin activity. It took 48 and 26 h, respectively, at 0 degree C for the two pools of prorenin at 37 degrees C to regain half of their maximum intrinsic activity at 0 degrees C. A direct immunoradiometric assay that detects active renin but not prorenin was able to detect cryoactivated prorenin. These results show that human prorenin can be reversibly cryoactivated in buffers of low ionic strength and has greater intrinsic activity at room temperature than at 37 degrees C.
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PMID:Reversible cryoactivation of recombinant human prorenin. 160 50

Human renin is synthesized as an inactive zymogen (prorenin) which is processed to the active form. We synthesized an 11-amino acid peptide which spans the human prorenin processing site in order to develop a simple assay to study human prorenin activation. Six enzymes which are capable of activating recombinant prorenin in vitro were studied. Four of these enzymes digested the synthetic peptide in a specific fashion, as analyzed by reverse-phase high-performance liquid chromatography. Amino acid analysis of the purified digestion products revealed that trypsin cleaves between Arg-Leu, the authentic processing site, while kallikrein, plasmin and elastase all cleaved at alternate sites. On the other hand, pepsin and cathepsin D did not cleave this substrate, suggesting that the activation of prorenin by these proteases might occur at a site distinct from the authentic processing site. Our data suggest that this synthetic peptide may be used as a simple and specific assay for prorenin activation.
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PMID:Characterization of prorenin activation using a synthetic peptide substrate. 165 85

To investigate the extrarenal release of renin, plasma active renin (PAR) and plasma trypsin activatable or inactive renin (PIR) in blood taken simultaneously from bilateral median cubital veins were measured in seven normal men before and after occlusion of the right upper arm with a standard cuff inflated halfway between systolic and diastolic blood pressures for 15 min. After 15 min, both PAR (P less than 0.05) with PIR (P less than 0.01) increased significantly in occluded arms compared with 0 min. PIR rose significantly (P less than 0.01) in occluded arms compared with controls at 15 min, but PAR was unchanged. The same studies were repeated after pretreatment with oral doses of either 1 mg prazosin or 10 mg propranolol at 1600 h and 2400 h on the previous day and 1 hour before the experiment. After prazosin, both PAR (P less than 0.05) and PIR (P less than 0.01) in occluded arms increased significantly at 15 min compared with those at 0 min and PIR rose significantly (P less than 0.01) in occluded arms compared with controls at 15 min, as observed in the control studies. On the other hand, after propranolol, the significant increment of PAR in the occluded arms was abolished. Moreover, the significant difference of PIR between the occluded and the control arms at 15 min was also abolished. These results indicate that inactive renin may be present in the vascular wall of the arm and released into circulation by a stimulus such as arm occlusion by mechanisms related to beta-adrenoceptors.
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PMID:Inactive renin release from the human arm vascular wall during upper arm occlusion: relation to beta-adrenoceptors. 166 64

To investigate the role plasma kallikrein plays in the in vivo activation of inactive renin, we measured plasma active renin, inactive renin, kallikrein and prekallikrein levels in 10 patients with disseminated intravascular coagulation (DIC), with 16 normal persons as controls. The plasma active renin concentration was expressed by the angiotensin I generation rate after the addition of sheep renin substrate. Plasma inactive renin was activated by trypsin. The plasma total kallikrein level was measured by an assay of kallikrein activity on synthetic substrate S-2302 after the addition of a prekallikrein activator. Plasma kallikrein was assayed by its activity on S-2302 without addition of the activator. The prekallikrein level was obtained by subtracting the kallikrein activity from the total kallikrein activity. A significant decrease in the plasma prekallikrein concentration was observed in DIC patients, as compared to that of controls (p less than 0.01). There was no significant difference in plasma levels of kallikrein, inactive renin, and the proportion of active renin between DIC patients and normal controls, but the active renin level was higher in DIC patients. There was no significant correlation between the level of plasma kallikrein and the proportion of active renin in either normal controls or DIC patients. These results are compatible with, but do not prove, the theory that plasma kallikrein plays a role in the in vivo activation of inactive renin.
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PMID:Plasma active renin, inactive renin and kallikrein in patients with disseminated intravascular coagulation. 168 Sep 81

There are changes in both the plasma renin system and plasma kallikrein system during parturition. To investigate the interrelationship between plasma inactive renin and plasma kallikrein, we measured plasma active renin, inactive renin, active kallikrein and inactive kallikrein in 21 parturient women just before delivery and on the 5th day after delivery, and also in 30 newborn babies upon birth and on the 5th day after birth. Plasma active renin was measured by radioimmunoassay of angiotensin I generated after the addition of an exogenous substrate. Inactive renin was activated by trypsin. Active kallikrein was measured by kallikrein activity on substrate S-2302. Inactive kallikrein was activated by an activator containing the Hageman factor and kininogen. The results showed a significant decrease in active renin, inactive renin, and a significant increase in active kallikrein, inactive kallikrein and the active/total kallikrein ratio in mothers on the 5th day after delivery. In vaginally delivered babies, a decrease in active renin and in the active/total renin ratio were observed on the 5th day after birth, but inactive renin, active and inactive kallikrein showed no change. In babies delivered by cesarean section, no change in either the renin or kallikrein level was found. The patterns of change in plasma active renin and inactive renin in mothers and babies are in keeping with previous suggestions that plasma inactive renin is prorenin. There was no correlation between the plasma active/total renin ratio or the plasma active kallikrein level in mothers and babies, either before or after delivery.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Plasma renin and kallikrein in parturient women and newborn babies. 168 Oct 14

To investigate the physiologic role of plasma inactive renin and its relationship to plasma kallikrein, we measured the changes in plasma active renin, inactive renin and prekallikrein levels in 14 uremic patients before and after hemodialysis. Blood was collected before, during and after hemodialysis, and prior to the next dialysis session. Plasma active renin was measured by radioimmunoassay of generated angiotensin I after addition of an exogenous substrate. Plasma inactive renin was activated by trypsin. Plasma prekallikrein was measured by the kallikrein-like activity on synthetic substrate S-2302 after activation of prekallikrein. The results showed that there was no change in blood pressure before, during or after dialysis, whereas the change in body weight after dialysis was significant. There was also no significant difference in the plasma active renin, inactive renin and prekallikrein levels for any of the collection periods. Plasma active renin was significantly correlated with inactive renin. The correlation between the active renin/total renin ratio and the plasma prekallikrein level was also not significant. These results suggest that in uremic patients undergoing chronic hemodialysis, the response of the renin system to acute plasma volume change is blunted. These data only provide evidence that plasma active renin is linked with inactive renin, but provide no evidence to support the idea that plasma inactive renin is a precursor of active renin or that plasma kallikrein is related to activation of inactive renin in vivo.
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PMID:Changes in plasma active and inactive renin and prekallikrein during hemodialysis. 168 91

Human renin is synthesized as a 406-amino acid preprorenin protein that is processed by a signal peptidase during secretion, to release prorenin as a 386-amino acid zymogen. The 46-amino acid "pro" domain is removed by a renin-processing enzyme, to produce enzymatically active renin, by cleavage at an Arg-Leu bond. The effects of the renin-processing enzyme can be mimicked by trypsin activation, where high concentrations of trypsin are incubated with prorenin for brief periods of time, followed by excess trypsin inhibitor to minimize secondary proteolytic processing by trypsin. In order to study the role of the pro segment in the secretion, folding, and activity of human renin, we engineered a construct where the pro domain from the preprorenin cDNA was deleted. This construct was introduced into mammalian cells and its expression was assayed in transient and stable systems. In COS-1 cells transfected with the prerenin expression vector pREN3, active renin was secreted with a specific activity of 1360 micrograms of angiotensin l/min/mg, compared with trypsin-activated prorenin, which has a specific activity of 818 micrograms of angiotensin l/min/mg. The active renin secreted in this system had a significantly reduced potency for the renin inhibitor SQ 32,970. These results demonstrate that the pro segment is dispensable for the folding and secretion of renin. A permanent cell line expressing the active form of renin was obtained by co-transfection of NRP cells with pREN3 and pHyg. A colony designated B/1 was identified, subcloned, and shown to secrete active renin (110 pg of renin/10(6) cells) optimally when maintained in both G418 and hygromycin.
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PMID:Stable expression, secretion, and characterization of active human renin in mammalian cells. 173 22

Previously, we unexpectedly observed that plasma inactive renin (trypsin-activatable renin) in bilaterally nephrectomized rats is not prorenin. To determine whether plasma inactive renin in anephric man is prorenin, we examined the immunological and biochemical properties of plasma inactive renin from five anephric patients. There were significant concentrations of inactive renin (5.33 +/- 2.08 ng/L.s) in plasma of anephric patients, while active renin was negligible (0.06 +/- 0.01 ng/L.s). The inactive renin from anephric patients could be immunoprecipitated 97 +/- 1% by specific antiserum against the prosegment portion of prorenin. Specific antimature renin serum completely inhibited the angiotensin-I-generating activity of inactive renin induced by trypsin treatment. The molecular mass of inactive renin from anephric patients (49.0 +/- 0 kDa), estimated by gel permeation high performance liquid chromatography, was similar to that of normal human plasma prorenin (48.2 +/- 0.8 kDa). These results indicate that plasma inactive renin in anephric man is prorenin, findings different from our previous observations obtained in anephric rats. Concanavalin-A chromatography separated inactive renin from anephric patients into three forms, including the column-unbound form, the loosely bound form, and the tightly bound form. Thus, in anephric man, differently glycosylated multiple forms of prorenin are released into the circulation from an extrarenal organ(s).
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PMID:Evidence for the presence of differently glycosylated forms of prorenin in the plasma of anephric man. 174 May 5

Isoelectric species of renin are physically heterogeneous. Recent evidence suggests that they may differ functionally, with some species producing natriuresis and diuresis, whereas others have no effect. A physiological function of secreted prorenin has not been documented in any species. The present study was designed to confirm and describe for the first time the renal effects of certain isoelectric species of prorenin. Anesthetized Sprague-Dawley rats were injected (0.1 ml) with trypsin-activated or nonactivated prorenin obtained from human ovarian follicular fluid. The dose chosen was calculated as sufficient to produce 2,300 ng angiotensin I.h-1.100 g rat body wt-1 in the presence of excess sheep substrate. Blood pressure, creatinine clearance, urine flow rate, and urine sodium, potassium, and osmolar excretion were measured. Activated prorenin from isoelectric peaks at isoelectric points (pI) 5.1, 5.2, 5.4, and 5.6 produced marked increases in urine volume (sixfold) and sodium excretion (7- to 10-fold) compared with the group receiving the vehicle (1% albumin in 0.9% saline). Activated prorenin from peaks at pI 4.9 and 5.8 produced no significant increase over the vehicle-only experiments. Captopril pretreatment (1 mg/kg iv) completely blocked the effects of peaks at pI 5.4 and 5.6. Interestingly, injection of nonactivated prorenin from peaks at pI 5.4 and 5.6 produced effects similar to the injection of activated prorenin from these peaks. Similarly, this effect was blocked by pretreatment with captopril. In summary, only certain isoelectric peaks of human prorenin whether activated, to active renin, or nonactivated produced a marked natriuresis and diuresis.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Differing renal effects of activated and nonactivated isoelectric peaks of human prorenin in rats. 175 May 22

Divergent conclusions exist as to whether inactive renin is present in nephrectomized rat plasma. A major factor contributing to this conflict may be related to significant changes in the "plasma blank" when trypsin-treated plasma is subjected to angiotensin I (AI) radioimmunoassay (RIA). In normal, but not nephrectomized rat plasma, AI-like substances are present in direct proportion to active renin. These substances are destroyed by trypsin. However, trypsin generates additional AI-like material, in both normal and nephrectomized rat plasma. This material, which is present in proportion to the renin substrate concentration, does not appear to be tetradecapeptide (TDP). In normal plasma, however, exogenous TDP is converted to AI in proportion to the active renin concentration and AI generation from TDP is increased by activation of inactive renin. However, in nephrectomized rat plasma, no AI generation from TDP was evident either before or after trypsin treatment. The coincident tryptic generation of a substance that quenches the levels of AI detected by RIA, combined with significant changes in the levels of endogenous and trypsin generated AI-like substances, may have significant bearing on the measured levels of inactive renin.
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PMID:Trypsin activation of inactive renin: plasma blanks and angiotensin I radioimmunoassay. 175 31


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