Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C1389183 (autodigestion)
 317 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Seven laboratories collaborating in a study of two intermediate purity plasminogen preparations (64/23, 63/6) observed that the amount of activator (urokinase or streptokinase) and the time of activation of plasminogen influenced the amount of plasmin generated. Using casein and a synthetic polypeptide (S-2251) as substrates, the authors subsequently showed that complete activation of plasminogen was difficult to achieve without acitivity losses due to plasmin autodigestion. Comparison of the polypeptide subunits (on SDS electrophoresis) of the various plasminogen activation mixtures with their plasmin activity allowed the conclusion that at maximum generation of plasmin from plasminogen, some plasminogen remains in the form of an inactive plasminogen intermediate (PLG-i).
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PMID:Activation of plasminogen as a feature in its assay. 14 Jan 14

Autocatalytic degradation of purified Bacillus subtilis neutral proteinase was examined under various conditions. At elevated temperatures, under non-inhibitory conditions, mature protein was rapidly degraded, but no accumulation of specific breakdown products occurred. However, by incubating purified neutral proteinase on ice during extended periods of time, specific peptides accumulated. These peptides were analysed by SDS/PAGE and Western blotting, and the N-terminal sequences were determined for the four major peptides, which had sizes of 30, 22, 20 and 15 kDa. Sequence data identified five fission sites in the neutral proteinase, three of which were identical with autodigestion target sites in thermolysin, a thermostable neutral proteinase. Comparison of the identified fission sites of the B. subtilis neutral proteinase with the known substrate-specificity of the enzyme indicated that they were in agreement, showing a preference for the generation of fissions at the N-terminal side of large hydrophobic residues, such as leucine, isoleucine and methionine. These results suggest a high degree of similarity in the three-dimensional structures of B. subtilis neutral proteinase and thermolysin.
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PMID:Identification of autodigestion target sites in Bacillus subtilis neutral proteinase. 212 7

1. A high-molecular-mass multicatalytic proteinase, ingensin, has been purified from rat liver and biochemically characterized. Trypsinization in the presence of ATP prevented the degradation of ingensin subunits. 2. Glutaraldehyde, which copolymerizes proteins, increased the apparent molecular mass of the subunits on SDS-PAGE, indicating the occurrence of covalent crosslinking of subunits. ATP, in this case, lowered the extent of covalent crosslinking. These results suggest that ATP altered the conformation of ingensin subunits. 3. Urea-induced autodigestion experiments demonstrated that some low-molecular-weight subunits selectively disappeared without changes in the contents of other subunits. The chymotryptic activity of the proteinase was more resistant to autodigestion than its tryptic activity. Therefore, we conclude that separate subunits of the enzyme are responsible for the different peptide-hydrolyzing activities.
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PMID:Molecular and biochemical properties of the ATP-stimulated multicatalytic proteinase, ingensin, from rat liver. 212 26

A purification procedure for neutral proteases from bacilli is described, in which bacitracin-silica was used as affinity medium. This enabled a one-step purification of the proteases directly from culture supernatant. Since neutral proteases are extremely sensitive towards autodigestion, conditions were chosen such, that autodigestion was largely prevented. Isopropanol appeared to be useful in both eluting the enzymes from the affinity medium, and inhibiting enzymatic activity during this step. The bacitracin-silica medium allowed high flow rates: with columns prepared for use in an FPLC system flow rates up to one column volume per minute were feasible, and still gave satisfactory results. The neutral proteases purified by this method were found to be homogeneous both by SDS-PAGE and analytical gel filtration.
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PMID:One-step affinity purification of Bacillus neutral proteases using bacitracin-silica. 249 14

Using components purified from human plasma, we have examined the effects of C1 inhibitor (C1 INH), the primary inhibitor of activated Hageman Factor (HFa) and Hageman factor fragment (HFf), on Hageman Factor (HF) autoactivation. When Hageman factor was exposed to a negatively charged surface, provided by either a glass cuvette or dextran sulfate, the addition of C1 INH gave a dose-dependent inhibition of the activity observed. The ability of C1 INH to decrease the maximal enzymatic activity generated was markedly temperature dependent with inhibition increasing as the temperature was raised from 4 degrees C to 37 degrees C. Although the rates of both autoactivation and inhibition were decreased at lower temperatures (4 degrees C), the latter rate was more sensitive to temperature modulation. When HF (final concentration 1 mumol/L) was incubated with C1 INH (0.54, 1.07, and 2.14 mumol/L) in the absence of an initiating surface, no increases in enzymatic activity were observed for up to 48 hours regardless of the C1 INH concentration. However, SDS polyacrylamide gel electrophoresis of the incubation mixture revealed that HF autodigestion had occurred by 48 hours despite the presence of C1 INH. In addition, the appearance of a new band suggested that a complex had been formed between the inhibitor and activated HF. Our findings indicate that C1 INH does not prevent HF autoactivation but rather inactivates the products of HF autodigestion.
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PMID:The effect of C1 inhibitor upon Hageman factor autoactivation. 348 56

Porcine pancreatic phospholipase A2 (PLA2) was immobilized to Sepharose 4B and porcine serum was passed through this affinity column. Bound substances were eluted by an EDTA-containing buffer and fractionated in a Sepharose 6B column. A single protein peak of the eluate from the latter column was found to inhibit PLA2 activity in a dose-dependent manner in an assay system using radioactive lecithin as a substrate and porcine pancreatic PLA2 as the enzyme source. The serum fraction containing the PLA2 inhibitory protein(s) (PIP) appeared inhomogeneous on SDS-polyacrylamide gel electrophoresis with two major bands close to each other, corresponding to a molecular weight of approximately 60,000. It was concluded that PIP might act as a protective principle against autodigestion in acute pancreatitis and other inflammatory diseases as well as playing a regulatory role in prostaglandin metabolism.
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PMID:Presence of a phospholipase A2 inhibitor in porcine serum. 652 Jan 25

DNA endonucleases in rat liver nuclei extracts were examined by SDS-polyacrylamide gel electrophoresis followed by zymogram analysis. Four polypeptides of 120, 54, 31 and 28 kDa, which have DNA endonuclease activity, were shown to occur in the extract isolated in the presence of phenylmethanesulfonyl fluoride (PMSF), a proteinase inhibitor. Isolation without PMSF, as well as storage at -20 degrees C, or autodigestion, resulted in multiplication of active polypeptides in the extracts. Trypsin digestion led to the appearance of an active > 140 kDa polypeptide, indicating the existence of a potential endonuclease precursor in the nuclear extract.
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PMID:In vitro proteolysis of endonucleases in rat liver nuclei extracts. 766 99

Ginger proteases in ginger rhizome (Zingiber officinale roscoe) were extracted from the ginger acetone powder and purified on DEAE-Sepharose and Sephadex G-75 columns. Before the purification, excess p-chloromercuribenzoate was added to the enzymes to prevent their autodigestion. The mercuribenzoate-proteases were further purified and fractionated by isoelectric focusing in Ampholine of pH 3-10 or pH 4-6. The proteases were fractionated into three components by the isoelectric focusing, having pI value of 4.5, 4.6 and 4.8 respectively. All these proteases had a molecular mass of 29,000 as measured by SDS-polyacrylamide gel electrophoresis and by TSK G2000SW XL gel chromatography. The Ampholine in the purified enzymes can quickly be removed by the gel chromatography of TSK G2000SW. Some divalent metal ions, such as Hg2+, Cu2+, Cd2+, and Zn2+, strongly inhibited these purified enzymes.
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PMID:Purification of ginger proteases by DEAE-Sepharose and isoelectric focusing. 787 61

We have developed a gel electrophoresis system that can concentrate proteins from spots cut out of up to 50 two-dimensional electrophoresis gels. During protein concentration, SDS is substituted with a non-ionic detergent (octyl beta-glucopyranoside) which allows digestion and MS analysis of the protein directly extracted from the gel without fixation or staining. The system avoids the problems associated with the digestion of dilute protein in multiple bands by (a) greatly reducing the gel volume for digestion and thus the amount of protease required, hence lowering contamination by autodigestion products, (b) reducing the volume of solvent required for extraction of protein from the gel, thus minimising loss of material to container surfaces, and (c) removing SDS which interferes with subsequent MS or HPLC analysis. The efficiency of protein recovery ranges between an average of 80% for proteins from silver stained two-dimensional gels to 90% for fluorescence and Coomassie-blue-stained gels. The method is compatible with MS analysis of very low amounts of protein from any staining system, but appears not to be useful for Edman sequencing of silver-stained or fluorescent-stained proteins since the amount of N-terminal blockage appears to increase as the amount of protein isolated from the two-dimensional gel decreases.
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PMID:Concentration of, and SDS removal from proteins isolated from multiple two-dimensional electrophoresis gels. 920 22

We evaluated the activation of mu-calpain in progesterone-activated human sperm. Semen collected from fertile donors with informed consent was liquefied and subjected to percoll gradient centrifugation. After exposure to different concentrations of progesterone, the samples were used for immunostaining, SDS-PAGE and Western blot analysis. An increase of the intracellular free calcium concentration in the sperm following the addition of progesterone was observed using fura-2 AM. Immunostaining using an antibody against active mu-calpain produced 6 distinct staining patterns: (1) the acrosome, (2) an equatorial segment, (3) the whole head, (4) the neck, (5) the neck and tail or (6) unstained sperm. After addition of progesterone, the predominant type changed from the neck type (90%) to the neck and tail type (79%). Western blot analysis using a pro-mu-calpain and a mu-calpain domain III antibody revealed autodigestion of mu-calpain, indicating activation by progesterone. Using calpain-specific inhibitors it was shown that calpain activation contributes to sperm motility as well as to the acrosome reaction. These results suggest the possibility that activation of mu-calpain in human sperm by progesterone plays an important role in fertilization.
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PMID:Role of calpain in human sperm activated by progesterone for fertilization. 1151 38


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