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)

Porcine and ovine 19-S thyroglobulins prepared from frozen glands in several buffers using slice extraction or homogenization, ammonium sulfate precipitation and DEAE-cellulose chromatography or Sepharose 6B gel filtration were contaminated with protease activity of pH optima 4.5 and 8.6, as shown by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Optimum temperatures of autodigestion were 37 degrees C at pH 4.5 and 25 degrees C at pH 8.6. Thyroglobulins prepared from unfrozen glands pH 7.2 in 0.1 M sodium phosphate using slice extraction, ammonium sulfate precipitation and Sepharose 6B gel filtration were devoid of acid proteolytic activity but still underwent autodigestion at pH 8.6. Diisopropylfluorophosphate was a potent inhibitor of the alkaline protease activity of ovine thyroglobulin preparations. In contrast to thyroglobulin obtained from frozen glands the proteins purified from fresh unfrozen glands at pH 7.2 only showed the 19-S and the 12-S species by electrophoresis in sodium dodecyl sulfate polyacrylamide gels. Very few bands migrating faster than 12-S were visible. After full reduction and S-alkylation of porcine and ovine thyroglobulins, no qualitative changes were observed in the gel electrophoresis pattern as compared to the unmodified proteins. Species of apparent mol. wt. corresponding to the native 12 S were the major component, strongly suggesting a mol. wt. of about 330 000 for the elementary peptide chains of pig and sheep thyroglobulins.
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PMID:Endogenous proteolytic activity and constituent polypeptide chains of sheep and pig 19 S thyroglobulin. 0 35

The level of adenosine 3':5'-cyclic monophosphate (cAMP) and the activity of adenyl cyclase were studied in the pancreas under normal conditions and during acute hemorrhagic pancreatitis induced by intraductal injection of fresh trypsin-bile-blood mixture. In addition, the adenyl cyclase was localized histochemically in the pancreas. Basal cAMP concentration and adenyl cyclase activity were 0.88 +/- 0.11 pmoles/mg wet tissue and 3.39 +/- 0.21 pmoles/mg protein/min, respectively. The acute pancreatitis drastically reduced the adenyl cyclase activity at 15 minutes to 1.66 +/- 0.54 pmoles/mg protein/min, and totally suppressed adenyl cyclase activity at 30 minutes after the onset of pancreatitis without affecting cAMP levels. The presence of sodium fluoride in the incubation medium prolonged the enzyme activity up to 45 minutes. The progressive disappearance of adenyl cyclase activity presumably resulted from the destruction of cellular integrity caused by autodigestion by the active proteolytic enzymes released during pancreatitis.
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PMID:Adenyl cyclase and cyclic AMP (cAMP) in acute experimental pancreatitis. 18 29

Limited chemical extraction of hydrophobic glycopeptides and subtotal autodigestion of the donor's cells and plasma membranes in undemineralized cortical bone in vitro reduces the putative quantity of haptenic substances absorbed by the recipient. Iodoacetic acid and sodium azide or other sulfhydryl group enzyme inhibitors added to the buffer solutions during in vitro autodigestion and estraction of intracellular alloantigens protects the bone matrix morphogenetic property against enzymatic degradation. The delayed hypersensitivity reaction induced by aseptically collected freeze-dried bone and the destruction of the bone morphogenetic property caused by radiation-sterilization is avoidable by sequential chemodigestion and chemosterilization of bone that preserves the maximum morphogenetic potential while transferring a minimum quantity of alloantigen.
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PMID:A chemosterilized antigen-extracted autodigested alloimplant for bone banks. 109 48

Rats fed a diet containing 2.5% sodium saccharin (NaSacc) displayed a rapid (24-36 hr) increase in tryptic and chymotryptic activity in the lower half of the small intestine and the cecum compared with control animals. Cecal pH of rats fed NaSacc was lower than controls. The effect of NaSacc on enzymatic activity of intestinal contents and on indigenous bacterial microflora was studied further in vitro. Intestinal contents incubated anaerobically with or without NaSacc revealed that the presence of NaSacc led to higher tryptic and chymotryptic activity and higher final pH. Changes in pH do not appear, however, to be important for the increased proteolytic activity induced by NaSacc since autodigestion of trypsin and chymotrypsin in filter-sterilized samples was only slightly affected by pH during in vitro incubation. Amylolytic activity, on the other hand, was stabilized by higher pH values. Saccharin stabilized chymotryptic and led to almost complete loss of amylolytic activity during incubation of filter-sterilized samples maintained at adjusted pH values. The amount of reducing sugars remaining in the NaSacc-containing contents from either cecum (in vivo) or from in vitro incubation of unsterilized small intestinal samples was greater than controls not containing NaSacc. The growth of six bacterial strains isolated from small intestinal contents and incubated in laboratory media was inhibited by NaSacc. Extracellular proteolytic activity from bacterial sources was undetectable after incubation of intestinal bacteria in laboratory media. The present results suggest that the effect of NaSacc upon digestive enzyme composition in the small intestine of rats is not mediated through a direct physiological effect of NaSacc on pancreatic exocrine secretion. It is hypothesized that an inhibition of enzymatic activity by NaSacc in the small intestine and the bacteriostatic effect of NaSacc on bacteria may be responsible for the increased proteolytic activity observed in vivo in the cecum following the feeding of a NaSacc-containing diet to rats.
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PMID:Effects of sodium saccharin on the activity of trypsin, chymotrypsin, and amylase and upon bacteria in small intestinal contents of rats. 257 3

Intrapancreatic activation of proteases is believed to play a major role in the pathogenesis of acute necrotizing pancreatitis. Several authors have questioned, however, the central role of trypsin in autodigestion of the pancreas. To clarify the direct effects of pancreatic enzymes and other related factors on acinar cells, we used the model of isolated pancreatic acini. Acini were prepared from male Wistar rats by collagenase digestion. Protein synthesis was measured by incubation of acini with [35S]methionine. Acini were resuspended thereafter in fresh buffer and further incubated for 30-90 min under various conditions [e.g., with pancreatic homogenates, ascites (from rats with pancreatitis induced by sodium taurocholate), pure pancreatic enzymes, and other factors]. The percentage of release of newly synthesized proteins into the culture medium was regarded as a biochemical parameter of cellular integrity. A morphologic score of cellular integrity was obtained via light microscopic evaluation of acini at the end of the various incubations by measuring the degree of cell lysis, loss of cell granules, ballooning, formation of vacuoles, and karyopyknosis. When normal [35S]methionine-labeled pancreatic acini were incubated with various factors, the percentage of release of labeled proteins into the medium was as follows: incubation with HEPES/Ringer's buffer, 1.8%; hemorrhagic pancreatic ascites, 3.8%; pancreatic homogenates, 2.0%; lipase, 1.8%; phospholipase A2, 3.0%; phospholipase A2 + lecithin, 3.2%; trypsin, 2.5%; 5% olive oil, 1.8%; ascites + olive oil, 78.3%; ascites + homogenized epididymal fat, 79.9%; lipase + olive oil, 32.0%; pancreatic homogenates + olive oil, 28.0%; diolein, 2.65%; and oleic acid, 62.9%. The cellular release of radiolabeled proteins showed an inverse correlation with cellular integrity as shown by light microscopy. We postulate that interstitial release of degradation products from triglycerides by lipase causes cellular disruption. Whereas phospholipase A2 and proteases do not seem to be very harmful in the early phases of cellular damage, lipase may play a major role in acute necrotizing pancreatitis.
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PMID:Role of pancreatic enzymes and their substrates in autodigestion of the pancreas. In vitro studies with isolated rat pancreatic acini. 291 45

The effects of biliary diversion on pancreatic enzyme activities of intestinal contents was studied in conscious rats prepared with biliary and pancreatic fistulae. Diversion of bile from the intestine for 1 day caused on 80% decrease in trypsin and chymotrypsin activities of intestinal contents, in spite of increased (230%) pancreatic trypsin and chymotrypsin secretion. Bile diversion in fed rats caused a smaller decrease (58%) in trypsin and chymotrypsin activities of intestinal contents. Sodium taurocholate (100 mumol/hr intraduodenally) partially reversed the changes in pancreatic secretion and intestinal contents' activities of trypsin and chymotrypsin caused by bile diversion. The results indicated that bile was important in controlling the rate of disappearance of trypsin and chymotrypsin activities from the small intestine. The mechanism for this was studied by comparing the rate of disappearance of trypsin activity in vivo and in vitro. Bovine trypsin, with or without sodium taurocholate, was infused intraduodenally into conscious rats deprived of bile-pancreatic juice and the recovery of trypsin activity from the small intestine determined. Taurocholate increased recovery of trypsin from the small intestine more than threefold, but inactivation of bovine trypsin in vitro was not retarded by sodium taurocholate. The results indicate that bile in the small intestine controls the rate of disappearance of intraluminal trypsin and chymotrypsin activities, probably by inhibiting their autodigestion in vivo. We previously reported that bile duct ligation in rats caused decreased trypsin and chymotrypsin activities in the small intestine, but increased pancreatic enzyme secretion. We concluded that trypsin and chymotrypsin underwent accelerated inactivation in the small intestine in the absence of bile. The present study was designed to explore the mechanism for the effects of bile deprivation on intraluminal proteolytic enzyme activities in the rat.
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PMID:Importance of bile in regulation of intraluminal proteolytic enzyme activities in the rat. 615 98

Autodigestion of two cysteine proteinases, calotropins DI and DII isolated from the latex of Calotropis gigantea, has been studied at pH 7.5 and 37 degrees C in the presence of an activating agent. Calotropin DI is more susceptible to autodigestion than calotropin DII. During autodigestion no interconversion of one calotropin to another has occurred, as verified by polyacrylamide gel electrophoresis in the presence and absence of sodium dodecyl sulfate. Immunologically, both calotropins are closely related, but they differ from papain and ficin. Both calotropins have blocked N-terminal amino acid residues. Their C-terminal amino acid sequences, determined by treatment with carboxypeptidase Y, are -(Pro, Ala)-Ala-Val-Tyr for calotropin DI and -(Ala, Val)-Ala-Pro-Tyr for calotropin DII. The tryptic peptide maps of their reduced and S-carboxymethylated derivatives suggest that both calotropins share a high proportion of common regions in their amino acid sequences. Calotropins DI and DII are two distinct proteinases, and they do not appear to be produced by autodigestion of a single precursor. Although they are inert to the common synthetic substrates of papain and ficin, their specificities toward oxidized insulin B chain are comparable to those of papain and ficin.
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PMID:Comparative studies on calotropins DI and DII from the latex of Calotropis gigantea. 643 Feb 36

Capillary electrophoresis (CE) and matrix assisted laser desorption time-of-flight mass spectrometry (MALDI-TOF-MS) were investigated as alternatives to sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis for peptide mapping with Staphylococcus aureus protease (V8) of a hydrophobic recombinant hepatitis C virus antigen, HC-31, which required 0.1% SDS for solubility. Controls (V8 only) or HC-31 digests were extracted with chloroform-methanol-water (1:4:3) to remove SDS, which interferes with MALDI-TOF, and high salt content, which affects CE. In two different runs by CE, the elution times of each of 11 peptide peaks were very reproducible (R.S.D. < 0.016). 25 fragments were resolved by MALDI-TOF-MS, including six smaller peptides (M(r) < 13 000) resulting from V8 autodigestion. MALDI-TOF-MS indicated that partial cleavages occurred, primarily at sites where there are paired glutamic and/or aspartic acid residues.
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PMID:Comparative peptide mapping of a hepatitis C viral recombinant protein by capillary electrophoresis and matrix-assisted laser desorption time-of-flight mass spectrometry. 884 66

A method for the isolation and primary monolayer culture of adult pig pancreatic endocrine (PE) cells was established. Cells were dissociated from the pancreas by autodigestion without addition of proteolytic enzymes and separated into distinct bands in a single centrifugation step using Histopaque-1077 (a mixture of polysucrose and sodium diatrizoate). The cells collected from an interfacial fraction were suspended in RPMI 1640 containing 11 mmol/l D-glucose with or without nicotinamide (0, 10, 20, 40 mmol/l), and then placed in culture dishes. Pancreatic cells formed a monolayer while fibroblasts became detached from the bottom of the dish when cultured in the presence of nicotinamide. More than 80% of monolayer-forming cells were stained for insulin, using an enzymatic method, and were identified as B-cells. Morphologically, the PE cells extended multiple processes terminating in growth-cone-like structures, as visualized by both light microscopy and scanning electron microscopy. Insulin secretion in response to glucose stimulation occurred for 35 days of incubation in the RPMI 1640 medium, with or without nicotinamide. Exposure of the cells to nicotinamide for 35 days resulted in a 2-3-fold increase in insulin secretion in response to high glucose stimulus (16.7 mmol/l) compared with low glucose (5.5 mmol/l). Glucose-induced Ca2+ responses were examined in individual cells cultured for 35 days in the presence of 10 mmol/l nicotinamide, using Ca2+ imaging with fura-2. These results indicate that it is possible to prepare pig PE cells in monolayer culture with low fibroblast contamination and to maintain functioning B-cells in vitro for relatively long periods. The present method provides useful preparations for further morphological and physiological studies on the differentiation, growth and regenerative capacity of islet cells.
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PMID:Establishment of monolayer culture of pig pancreatic endocrine cells by use of nicotinamide. 988 27

Chymotrypsin is a prominent member of the family of serine proteases. The present studies demonstrate the presence of a native fragment containing 14 residues from Ile16 to Trp29 in alpha-chymotrypsin that binds to chymotrypsin at the active site with an exceptionally high affinity of 2.7 +/- 0.3 x 10(-11) M and thus works as a highly potent competitive inhibitor. The commercially available alpha-chymotrypsin was processed through a three phase partitioning system (TPP). The treated enzyme showed considerably enhanced activity. The 14 residue fragment was produced by autodigestion of a TPP-treated alpha-chymotrypsin during a long crystallization process that lasted more than four months. The treated enzyme was purified and kept for crystallization using vapour the diffusion method at 295 K. Twenty milligrams of lyophilized protein were dissolved in 1 mL of 25 mM sodium acetate buffer, pH 4.8. It was equilibrated against the same buffer containing 1.2 M ammonium sulfate. The rectangular crystals of small dimensions of 0.24 x 0.15 x 0.10 mm(3) were obtained. The X-ray intensity data were collected at 2.2 angstroms resolution and the structure was refined to an R-factor of 0.192. An extra electron density was observed at the binding site of alpha-chymotrypsin, which was readily interpreted as a 14 residue fragment of alpha-chymotrypsin corresponding to Ile-Val-Asn-Gly-Glu-Glu-Ala-Val-Pro-Gly-Ser-Trp-Pro-Trp(16-29). The electron density for the eight residues of the C-terminus, i.e. Ala22-Trp29, which were completely buried in the binding cleft of the enzyme, was of excellent quality and all the side chains of these eight residues were clearly modeled into it. However, the remaining six residues from the N-terminus, Ile16-Glu21 were poorly defined although the backbone density was good. There was a continuous electron density at 3.0 sigma between the active site Ser195 Ogamma and the carbonyl carbon atom of Trp29 of the fragment. The final refined coordinates showed a distance of 1.35 angstroms between Ser195 Ogamma and Trp29 C indicating the presence of a covalent linkage between the enzyme and the native fragment. This meant that the enzyme formed an acyl intermediate with the autodigested fragment Ile16-Trp29. In addition to the O-C covalent bond, there were several hydrogen bonds and hydrophobic interactions between the enzyme and the native fragment. The fragment showed a high complementarity with the binding site of alpha-chymotrypsin and the buried part of the fragment matched excellently with the corresponding buried part of Turkey ovomucoid inhibitor of alpha-chymotrypsin.
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PMID:Detection of native peptides as potent inhibitors of enzymes. Crystal structure of the complex formed between treated bovine alpha-chymotrypsin and an autocatalytically produced fragment, IIe-Val-Asn-Gly-Glu-Glu-Ala-Val-Pro-Gly-Ser-Trp-Pro-Trp, at 2.2 angstroms resolution. 1565 93


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