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

---

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.
...
PMID:Reversible cryoactivation of recombinant human prorenin. 160 50

In this paper we describe a routine method of measuring inactive renin in rat plasma. The activation was performed by trypsin, at optimal concentration and incubation conditions. The trypsin treatment formed an interfering and high-performance liquid chromatography-verified tetradecapeptide-like material, which was removed before the assay by a simple batchwise use of a cation-exchange resin. The concentration of activated inactive renin was measured by an antibody-trapping method after the addition of exogenous angiotensinogen. Angiotensinogen was added in order to compensate for the trypsin destruction of angiotensinogen and in order to measure the parameter of renin concentration. The inactive renin concentration in plasma of conscious male rats was 0.48 +/- 0.13 Goldblatt units (GU) per litre (n = 38). This corresponds to 66% (range 42-92%) of the total renin concentration. Physiological experiments in conscious rats were initiated, demonstrating that nephrectomy decreased the inactive renin concentration from 0.45 +/- 0.14 to 0.27 +/- 0.05 GU/l after 24 h (n = 21; P less than 0.01). Submandibular sialoadenectomy decreased the plasma inactive renin concentration from 0.45 +/- 0.11 to 0.34 +/- 0.06 GU/l (n = 12; P less than 0.05) after 7 days. Combined sialoadenectomy and nephrectomy decreased the plasma inactive renin concentration from 0.45 +/- 0.11 to 0.24 +/- 0.06 (n = 12; P less than 0.01).
...
PMID:Measurement of inactive renin in rat plasma: effect of nephrectomy and sialoadenectomy on the plasma concentration. 216 Apr 91

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 inactive renin from chorion laeve was completely inactive and did not bind to pepstatin-aminohexyl-Sepharose; however, that from chorion culture medium was partially active and completely bound to the pepstatin gel, indicating that each molecule is partially activated. Trypsin-activated inactive renins from both sources were identical with human renal renin in terms of pH optimum and Km. Specific activities of trypsin-activated inactive renin from chorion laeve and chorion culture medium were 529 Goldblatt units/mg of protein and 449 Goldblatt units/mg of protein, respectively. Amino acid sequence analysis of both of the purified inactive renin preparations demonstrated a leucine residue at the amino terminus. The sequence of 11 additional amino acids was identical in both and agreed with that predicted from the base sequence of the renin gene. These findings indicate that preprorenin is converted to prorenin following removal of a 23-amino acid signal peptide and that the native inactive renin, whose amino acid sequence commences with Leu-Pro-Thr..., is prorenin.
...
PMID:Pure human inactive renin. Evidence that native inactive renin is prorenin. 267 Sep 24

We measured the levels of trypsin-releasable spasmogenic substances (TRSS) in the plasma of spontaneously hypertensive rats (SHR) during the development of hypertension. TRSS levels (means +/- SEM, N = 4) were significantly higher at 12 weeks (7.13 +/- 1.05 micrograms bradykinin equivalents (BKE)/ml plasma) and 24 weeks (6.87 +/- 0.60 micrograms BKE/ml) compared to 8 weeks (3.3 +/- 0.55 micrograms BKE/ml) and to normotensive Wistar Kyoto (WKN) rats, whose levels were 3.74 +/- 0.74 micrograms BKE/ml at 24 weeks and did not change significantly during the period studied. The mean arterial pressure (MAP) of SHR was 150-170, 160-180 and 170-220 mmHg at 8, 12 and 24 weeks, respectively, whereas the WKN MAP was 110-120 mmHg at 24 weeks. The increase in total TRSS was due to substances which elicit the slow contraction of the isolated guinea pig ileum and which could be distinguished from BK, T-kinin and other BK homologues by gel filtration on Sephadex G-25, gradient elution chromatography on CM-cellulose and by the slow rate of contraction of the guinea pig ileum. All of these properties are the same as those we have previously demonstrated for TRSS of Goldblatt 1-kidney 1-clip renal hypertensive rats and which are due, at least in part, to a 14 amino acid peptide whose composition does not correspond to any known spasmogenic substance.
...
PMID:New trypsin-releasable spasmogenic substances in the plasma of spontaneously hypertensive rats. 322 26

Native human renin, produced from the culture of human chorionic trophoblasts, has been purified to homogeneity on a milligram scale using a five-step purification scheme. The chorion cells secrete 50-200 milliGoldblatt Units of trypsin-activatable prorenin per ml into the medium. The pro-enzyme is partially purified by ammonium sulfate fractionation and chromatographies on QAE-Sephadex and cibracon blue-agarose. Following conversion of prorenin to the active enzyme by porcine trypsin, the renin is purified to homogeneity by affinity chromatography and gel filtration. Chorionic prorenin has a molecular weight of 43,000; the active enzyme 40,000. Both proteins exist as a single polypeptide chain as determined by SDS-polyacrylamide gel electrophoresis under reducing conditions. The average specific activity of six different preparations was found to be 1072 Goldblatt Units/mg. The amino acid composition and N-terminal sequence of the active enzyme has been determined and is identical to the human kidney enzyme. Microheterogeneity of chorionic renin was demonstrated by isoelectrofocusing analysis. The physical characterization of chorionic renin is compared with that reported for the human kidney enzyme.
...
PMID:Human placental chorionic renin: production, purification and characterization. 328 35

Assay of inactive renin in unfractionated mouse plasma is difficult and often impossible because of high concentrations of active renin and plasma protease inhibitors. Therefore, 0.025 to 0.05 ml of plasma or amniotic fluid from mice was separated by high performance liquid chromatography (HPLC) using a silica-based size exclusion column. The eluates were examined for enzymatically active renin before and after limited proteolysis with trypsin. Since inactive renin eluted as a single peak corresponding to a molecular weight of 38,000 daltons and the elution of active renin was markedly retarded, inactive and active renin were partially separated. compared with plasma, inactive renin in amniotic fluid eluted as a broader and sometimes diphasic peak, suggesting heterogenicity. The rapid and reliable separation by HPLC provided a more than 300-fold purification of inactive renin. Despite low concentrations of plasma protease inhibitors, a 1 000 000-fold molar excess of trypsin (1 mg/ml) was needed for optimal activation. The necessity for high trypsin concentrations for activation may partly be explained by enzyme kinetic considerations. By combining HPLC with trypsin activation, inactive renin was readily measured and found to be 9.4 (5.1-13.2) GU (Goldblatt units)/l in normal and 8.0 (5.1-12.2) GU/l in sialoadenectomized and nephrectomized mouse plasma, which is higher than previously determined in our laboratory. The concentration was 12.4 (8.8-16.1) GU/l in amniotic fluid. Thus, the concentration of inactive renin in plasma is almost as high as active renin in normal mice (17.6 GU/l) and is uninfluenced by the decrease of active renin to 0.3 GU/l after sialoadenectomy and nephrectomy.
...
PMID:Quantitative activation and determination of inactive renin by high performance liquid chromatography. 329 36

Renin biosynthesis was studied in a juxtaglomerular cell tumor. The tumoral tissue had a high renin content (180 Goldblatt Units/g of tissue), was heavily stained by immunofluorescence using human renin antiserum, and exhibited numerous characteristic secretory granules by electron microscopy. In one series of experiments, renin biosynthesis was studied in tissue slices, by following the incorporation of radiolabeled amino acids into specific immunoprecipitable renin. Time course studies showed that renin was first synthesized in a high molecular weight form, 55,000 mol wt, i.e., 10,000 mol wt higher than that of active renin, and was then converted into a 44,000-mol wt form. In a second series of experiments renin tumoral cells were cultured. Small, round, birefringent cells obtained after collagenase digestion produced renin in both primary culture and subculture media. After 5 d most of the renin found in the culture medium was inactive, but could be activated by trypsin treatment. The tumoral tissue exhibited a strong renin immunofluorescence and numerous secretory granules were observed by electron microscopy. In contrast, the renin-producing cells isolated from this tumor and grown in culture showed little renin immunofluorescence and no secretory granule could be observed. The renin-producing cells in primary culture and subculture were pulsed with radiolabeled amino acids, and immunoprecipitable radiolabeled renin was found in the culture media, thus demonstrating the actual biosynthesis of the enzyme. This renin was not stored inside cultured cells but was rapidly released into the medium and had a molecular weight of 55,000. No conversion of this inactive high molecular weight renin into the active, 44,000 mol wt form of renin was observed. We postulate the existence of two pathways for the processing, packaging, and secretion of renin in the tumoral cells: in juxtaglomerular cells of tumoral tissue renin is synthesized as a preprorenin and rapidly converted into prorenin (55,000 mol wt), which is in turn packaged in secretory granules where it is processed into active renin (44,000 mol wt) and finally secreted; in the cultured tumoral cells renin is still biosynthesized as a preprorenin molecule and then converted into prorenin, but is neither stored as granules nor processed into active renin. In this case the renin is released in an inactive form.
...
PMID:Renin biosynthesis by human tumoral juxtaglomerular cells. Evidences for a renin precursor. 632 35

The presence of renin in parasympathetically elicited mouse saliva was demonstrated by using both the antibody trapping method, which measures renin's enzymatic activity, and a direct radioimmunoassay, which detects the renin molecule by its antigenic properties. Crossed immunoelectrophoresis of saliva samples using an antiserum elicited against pure submaxillary renin showed only one precipitation line, indicating the presence of only one form of renin. The position of the line was similar to that found when submaxillary gland extract was subjected to crossed immunoelectrophoresis. Tandem crossed immunoelectrophoresis showed complete identity between antigenic determinants in submaxillary and salivary renin. An apparent molecular weight of about 35 000 and 38 000 was found when saliva samples were subjected to gel filtration on Ultrogel AcA 44 and Sephadex G-100, respectively. No high molecular weight forms were present and no inactive forms could be demonstrated after limited pepsin or trypsin proteolysis. The specific enzymatic activity of renin in pilocarpine saliva was 0.37 Goldblatt Units (G.U.) . microgram-1, which is identical to that of pure submaxillary gland renin (0.41 G.U. . microgram-1) and to that to the storage form of renin in the submaxillary gland (0.4 G.U. . microgram-1) An identical Km value was found for salivary renin, 1.01 microM, and for pure submaxillary renin, 0.98 microM. It is concluded that renin in pilocarpine-elicited saliva is similar to the storage form of renin in the submaxillary gland with respect to molecular weight, enzymatic and immunological properties.
...
PMID:High levels of active 40 000-dalton renin in mouse saliva, but no evidence of inactive or high molecular weight forms. 702 27

1. Biochemical characteristics of a renin-like enzyme secreted by a pulmonary adenocarcinoma have been studied. A very high renin content was revealed by both enzymatic (10.4 Goldblatt units/g of tissue) and direct radioimmunoassay of immunoreactive renin (23 Goldblatt units/g of tissue). 2. The higher value by direct radioimmunoassay suggested the presence of an inactive form of the enzyme. Indeed 40--100% activation occurred with treatment by trypsin or pepsin or prolonged dialysis at pH 7.4 with or without prior acid dialysis. This neutral activation was completely abolished by a serine proteinase inhibitor. 3. A large fraction of the renin in this tumour is inactive. In comparison with other prohormones produced in tumours the findings support strongly the proposition that renin passes through a proenzyme step in synthesis.
...
PMID:Activation of renin in an anaplastic pulmonary adenocarcinoma. 703 19

Biochemical and immunological characteristics of renin secreted by two malignant renin-secreting tumors [pulmonary (PT) and paraovarian (POT)] were studied. They both contain inactive renin (IR), as renin activity of tumoral extracts was able to be increased after acid activation or trypsin treatment (10.1 to 20.8 Goldblatt units/g tissue for PT and 1.4 to 3.71 for POT). Renin activity after activation reached the value obtained by direct RIA of human renin (23 and 3.4, respectively), as both forms are recognized by renin antiserum. Both enzymatic activities could be completely inhibited by renin antiserum. Displacement curves for the two tumoral renins paralleled the MRC renin in the direct RIA. After chromatography on affigel blue, active renin was not bound to the gel, and inactive renin eluted only with 1 M NaCl. On pepstatin A Sepharose and CBL-pepstatin Sepharose (an N-modified-pepstatin), a separation of the two forms of pulmonary renin was obtained; inactive renin eluted with breakthrough proteins, whereas active renin was strongly bound to the gel. After this affinity chromatography, the molecular weights of inactive and active renin, determined on Ultrogel, were very close (46,000 and 42,500). We conclude that 1) ectopic renin in these cases in similar to the renal enzyme; 2) renin can be secreted in an inactive form, supporting the hypothesis of an inactive initial state of renin; and 3) molecular weight differences between the two forms are very slight.
...
PMID:Biochemical and immunological characterization of ectopic tumoral renin. 703 66


1 2 Next >>

---