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Query: UNIPROT:P00790 (PGA)
 2,475 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Cobalamin (Cbl; vitamin B(12)) malabsorption in pancreatic insufficiency can be partially corrected by bicarbonate and completely corrected by pancreatic proteases but the mechanisms involved are unknown. Because saliva contains enough R-type Cbl-binding protein (R protein) to bind all of the dietary and biliary Cbl, it is possible that R protein acts as an inhibitor of Cbl absorption and that pancreatic proteases are required to alter R protein and prevent such inhibition. To test this hypothesis we studied the ability of R protein and intrinsic factor (IF) to compete for Cbl binding and ability of pancreatic proteases to alter this competition. Human salivary R protein bound Cbl with affinities that were 50- and 3-fold higher than those of human IF at pH 2 and 8, respectively. Cbl bound to IF was transferred to an equal amount of R protein with t((1/2))'s of 2 and 90 min at pH 2 and 8, respectively, and within several hours respective ratios of R protein-Cbl/IF-Cbl of 50 and 2 were observed. Cbl bound to R protein was not transferred to IF at either pH 2 or 8. Incubation of R protein with pancreatic proteases at pH 8 led to a 150-fold decrease in its affinity for Cbl. Incubation of R protein-Cbl with pancreatic proteases led to complete transfer of Cbl to IF within 10 min. Gel filtration studies with R protein-[(57)Co]Cbl and (125)I-R protein showed that pancreatic proteases partially degraded R protein. Pancreatic proteases differed in their ability to effect these changes with trypsin > chymotrypsin > elastase. Pancreatic proteases did not alter IF in any of the parameters mentioned above. Pepsin failed to alter either R protein or IF. THESE STUDIES SUGGEST THE FOLLOWING: (a) that Cbl is bound almost exclusively to R protein in the acid milieu of the stomach, rather than to IF as has been assumed previously; (b) that Cbl remains bound to R protein in the slightly alkaline environment of the intestine until pancreatic proteases partially degrade R protein and enable Cbl to become bound exclusively to IF; and (c) that the primary defect in Cbl absorption in pancreatic insufficiency is a lack of pancreatic proteases and a failure to alter R protein and effect the transfer of Cbl to IF. These studies also suggest that the partial correction of Cbl malabsorption observed with bicarbonate is due to neutralization of gastric HCl, since at slightly alkaline, pH IF can partially compete with R protein for the initial binding and retention of Cbl.
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PMID:Effect of proteolytic enzymes on the binding of cobalamin to R protein and intrinsic factor. In vitro evidence that a failure to partially degrade R protein is responsible for cobalamin malabsorption in pancreatic insufficiency. 2 56

The action of six different enzymes on the function and structure of Factor H was investigated by use of sodium dodecyl sulphate/polyacrylamide-gel electrophoresis, haemagglutination, two enzyme-linked immunosorbent assay systems and an assay for Factor I cofactor activity. Six monoclonal antibodies directed against the 38 kDa tryptic fragment of Factor H [which contains the binding site for C3b (a 180 kDa fragment of the third component of complement) and the cofactor activity] were also used to detect cleavage products derived from the same fragment. Elastase, chymotrypsin A4 or trypsin first cleaved Factor H to 36-38 kDa fragments carrying all six monoclonal anti-(Factor H)-binding sites. In parallel, the interaction of Factor H with surface-bound C3b was lost, whereas the cofactor function was preserved. Further cleavage of the 36-38 kDa fragments into two 13-19 kDa fragments (one carrying the MAH4 and MRC OX 24 epitopes, the other the MAH1, MAH2, MAH3 and MRC OX 23 epitopes) destroyed cofactor activity. Pepsin, bromelain or papain rapidly split off a 13-15 kDa fragment of Factor H carrying the MAH1, MAH2, MAH3 and MRC OX 23 epitopes and destroyed all tested functions of Factor H. Ficin cleaved Factor H into disulphide-linked fragments smaller than 25 kDa, but did not affect the functions of the Factor H molecule. The 38 kDa tryptic fragment of Factor H is the N-terminal end of the Factor H molecule, as determined by N-terminal sequence analysis. A model is presented of the substructure of Factor H.
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PMID:Structural and functional analysis of the complement component factor H with the use of different enzymes and monoclonal antibodies to factor H. 293 33

Apolipoprotein AI of human high-density lipoproteins is secreted by hepatocytes as a proapolipoprotein with a N-terminal hexapeptide sequence (Arg-His-Phe-Trp-Gln-Gln-) which differs from the prosequence of rat apolipoprotein AI (Trp-Asp-Phe-Trp-Gln-Gln). The two proteins have in common the unusual cleavage site -Gln-Gln-Asp-Glu-. It is hydrolysed by a specific serum proteinase with the release of mature apo AI. We synthesized a model substrate for the study of the final processing of pro-apo AI by the serum proteinase. It is an undecapeptide embracing the human pro-hexapeptide sequence and the first five N-terminal residues of apo AI, covalently linked to a hydrophilic resin. The N-terminal arginine residue was 3H-labelled. [formula; see text] This sequence was not cleaved by human serum under the conditions under which rat serum processes the pro-form of apo AI secreted by rat hepatocytes. Pepsin and chymotrypsin fragmented the undecapeptide at sites characteristic for these proteinases. We conclude that the proteolytic cleavage at the specific site (-Gln-Gln-Asp-Glu-) requires the correct conformation in addition to the specific amino-acid sequence.
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PMID:Processing of proapolipoprotein AI requires specific conformation. 392 Oct 42

The primary structure of rat liver microsomal glutathione transferase has been determined. The 14C-carboxymethylated protein was fragmented with CNBr and proteolytic enzymes. The basis of the analysis was information from sequenator degradations of the intact protein, the largest CNBr fragment, and a large COOH-terminal fragment derived from a digest with Glu-specific staphylococcal protease. Remaining, smaller fragments were analyzed with the manual dimethylaminoazobenzene isothiocyanate method. Pepsin and limited acid hydrolysis were used to obtain peptides to confirm and overlap hydrophobic structures in the COOH-terminal half of the protein where trypsin and chymotrypsin failed to give any cleavage. Combined, these data permit the deduction of a 154-residue amino acid sequence. No evidence for micro-heterogeneity was obtained. The NH2-terminal alanine residue has a free alpha-amino group and the cysteine residue involved in activation of the enzymatic activity by sulfhydryl reagents is at position 49. The protein chain contains three regions with predictions for long beta strand secondary structures (positions 11-26, 103-120, and 131-145). Predictions may be inaccurate in membrane-associated proteins, but two of these regions also affect the three most hydrophobic segments. Thus, residues 11-35 form a long, largely hydrophobic part interrupted by only one charged residue (Lys-25), and residues 81-97 and 114-126 constitute the most hydrophobic segments directly noticeable from the hydrophilicity curve of the protein chain. These special parts of the molecule are of interest in relation to membrane interactions.
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PMID:Microsomal glutathione transferase. Primary structure. 393 48

1. With the aid of a coupled system involving glutathione reductase, the reaction of glutathione with the disulphide bonds of purified proteins has been studied. 2. Bovine serum albumin, conalbumin, lysozyme, trypsin inhibitors from egg white, lima bean and soya bean either did not react with glutathione or reacted only slightly. With these proteins reactivity was markedly increased by limited proteolysis. 3. Bovine and human gamma-globulins, fibrinogen and beta-lactoglobulin exhibited some reactivity (less than 15%) with glutathione and again this was increased by limited proteolysis. Pepsin, trypsin and chymotrypsin exhibited greater reactivity than the proteins previously mentioned. Di-isopropylphosphoryl-chymotrypsin exhibited less reactivity than chymotrypsin, suggesting that autolysis under the experimental conditions used contributed towards the reactivity of this protein. Proteolysis also increased the reactivity of these proteins. The three disulphide bonds of insulin were reduced by glutathione. 4. Above 35 degrees the disulphide bonds of serum albumin show a progressive increase in reactivity and at 55 degrees half of the bonds become accessible to glutathione. 5. From the results obtained with the proteins investigated, the conclusion reached is that the disulphide bonds of native proteins are structurally protected and do not react with glutathione under physiological conditions.
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PMID:The reactivity of the disulphide bonds of purified proteins in relationship to primary structure. 486 Apr 70

The flagellar complex of the unusual motile spermatozoon of the fungus gnat, Rhynchosciara sp, does not conform to the usual "9 + 2" filament pattern but rather consists of over 350 pairs of filaments (doublet microtubules) distributed in a spiral array. Experiments were designed to disrupt and extract flagellar microtubular components from spermatozoa of the fungus gnat. Pepsin, chymotrypsin, potassium iodide, urea, and heat were used to extract specific portions of microtubule walls Such experiments provide information on the composition of the wall and the existence of wall sites selectively sensitive to various treatments Results obtained include: (a) doublet microtubules are comprised at least in part of protein, and all subunits are probably not identical; (b) a portion of the B subfiber is apparently more sensitive to disruption than other portions of the doublet microtubule; and (c) the ac cessory singlet microtubules may be chemically different from the doublet microtubules
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PMID:Electron microscope studies of spermatozoa of Rhynchosciara Sp. I. Disruption of microtubules by various treatments. 504 61

Carbenoxolone may bind to enzymes, inhibiting or activating them. Enzymes inhibited are human pepsins 1, 3 and 5, human pepsinogens 1, 3 and 5, swine pepsin, bovine trypsin, bovine chymotrypsin, porcine elastase, human gastric proteinase 2, human gastric prostaglandin 15-OH dehydrogenase and delta-5 reductase, and pronase. Enzymes activated are papain, bovine carboxypeptidase and gastric microsomal glycosyl transferase. Enzymes unaffected are human pancreatic amylase and porcine pancreatic lipase. Binding occurs away from the active site; inhibition thus occurs when binding impedes access of substrate to, or products from, the active site, and activation when access is facilitated. Carbenoxolone causes increased secretion of mucus; this action can be explained by activation of the gastric glycosyl transferases. Carbenoxolone also causes intraluminal loss of peptic activity and diminished secretion of pepsins; these actions are explained respectively by intraluminal inhibition of the pepsins and intramucosal inactivation of the pepsinogens, particularly of the peptic ulcer-associated enzyme, pepsin 1. The healing effect of carbenoxolone in peptic ulcer involves these actions together with a reduced turnover of gastric mucosal cells. Pepsins 1 and 3 have collagenolytic activity, causing release of alpha-chains from native collagens. Pepsin 1 is five-fold the more active. Carbenoxolone inhibits peptic collagenolysis.
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PMID:The actions of carbenoxolone on enzymes and their relation to its therapeutic effect. 678 51

The effects of various proteolytic enzymes on the high molecular weight protein (connectin) present in a direct sodium dodecyl sulfate extract of myofibrils from chicken breast muscle were studied in detail. To keep the high molecular weight proteins intact, myofibrils had to be prepared in the presence of EGTA. Trypsin, chymotrypsin, papain, and nagarse readily hydrolyzed connectin (doublet band of titin) and the band 3 protein (N2-line protein). Pepsin did not attack connectin, but digested the band 3 protein and myosin. Calcium-activated neutral proteinase hydrolyzed the band 3 protein, leaving connectin intact. On the other hand, serine protease digested connectin but not the band 3 protein.
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PMID:Connectin, an elastic protein of muscle. Effects of proteolytic enzymes in situ. 702 43

1. Pigs (sixteen/diet) were weaned at 2 d of age and given liquid diets (200 g dry matter/l) during a 26 d experiment. The pigs were fed on a scale based on live weight. Dried skim-milk was the only source of protein in diet U and was partially or totally replaced by a soya-bean isolate (diet B) or a concentrate (diets C and D). Soya-bean protein provided 500, 700 or 350 g/kg total crude protein (nitrogen x 6.25) in diets B, C and D respectively. 2. Performance was similar for diets B and D, but poorer than that of pigs given diet U. The apparent digestibility and retention of N of these diets was similar. Pigs given diet C scoured severely and twelve died. 3. Protein digestion was studied in pigs given diets U, B and D, killed at 28 d of age, at the termination of the feeding experiment. The dry matter content and proportion of N in the digesta in the stomach were reduced in pigs given soya-bean protein. Pepsin concentrations in digesta and stomach tissue were unchanged. 4. The concentrations of trypsin and chymotrypsin in the pancreas were greater in pigs given the soya-bean protein concentrate compared with milk protein, but only the increase in trypsin was significant (P less than 0.01). Digesta from the small intestine of pigs given the soya-bean-protein isolate contained less chymotrypsin (P less than 0.05). There were no differences in the proportion of non-protein-N in the total N in the digesta, suggesting that proteolysis of the milk and soya-bean proteins were equally by 28 d of age.
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PMID:Artificial rearing of pigs. 12. Effect of replacement of dried skim-milk by either a soya-protein isolate or concentrate on the performance of the pigs and digestion of protein. 720 50

Matrix vesicles (MV), microstructures which rapidly accumulate Ca2+ and induce mineral formation in vitro, are linked to type II and X collagens and proteoglycans in the hypertrophic cartilage. However, the roles of these matrix proteins on MV function are not known. This led us to investigate the influence of type II and X collagen binding on Ca2+ uptake by MV. MV isolated from chicken growth plate cartilage were treated with pure bacterial collagenase and 1 M NaCl in synthetic cartilage lymph to selectively and completely remove associated type II and X collagens. Uptake of 45Ca2+ by these collagen-depleted vesicles was markedly reduced. Further treatment with detergent, which disrupted the membrane, restored Ca2+ uptake, indicating that the vesicle membrane structure and the nucleational core inside the vesicle lumen were still intact after the collagenase and 1 M NaCl treatments. Readdition of either native type II or X collagen to the collagenase, 1 M NaCl-treated MV stimulated their Ca2+ uptake to levels similar to those of untreated vesicles. Pepsin-treated type II and X collagens were less effective in stimulating Ca2+ uptake, indicating that non-triple helical domains of these collagens were involved. The pepsin treatment of these collagens also decreased their binding to annexin V (anchorin CII), one of three annexins found in MV, suggesting that annexin V is involved in mediating the binding of type II and X collagens to the MV surface. Furthermore, treatment of collagenase, 1 M NaCl-treated MV with chymotrypsin, which damaged annexin V as well as many other MV proteins, prevented the stimulation of Ca2+ uptake by these collagens. Thus, the interaction between type II and X collagens with MV activates the influx of Ca2+ into MV and may play an important role in calcification of the vesicles.
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PMID:Stimulation of calcification of growth plate cartilage matrix vesicles by binding to type II and X collagens. 815 77


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