Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P00790 (PGA)
 2,475 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Collagen was isolated by acetic acid extraction in the presence of protease inhibitors and also by pepsin digestion from the skins of dogs affected with the Ehlers-Danlos syndrome and the skins on non-affected dogs. The collagen preparations isolated by acetic acid extraction from the Ehlers-Danlos syndrome-affected dog skin contained a greater proportion of alpha-chains than the collagen preparations from the normal dog skin. When the collagen from the Ehlers-Danlos syndrome-affected dog skin was reduced with NaBH4 before heat denaturation, and electrophoresis, there was a greater proportion of beta-chains present. The collagen isolated from the normal dog skin was not affected by the NaBH4 reduction. Collagen preparations isolated by pepsin digestion from both the Ehlers-Danlos syndrome-affected dog skin and the non-affected dog skin contained the same quantity of alpha- and beta-chains. In addition, collagen from both affected and non-affected dog skins isolated by pepsin digestion contained 10-11% type III collagen as determined by the interrupted sodium dodecyl sulfate polyacrylamide gel electrophoresis method. Pepsin digestion of the collagens isolated by acetic acid extraction in the presence of protease inhibitors from the skins of affected and non-affected dogs eliminated the differences between the alpha:beta ratios of the affected and non-affected collagen preparations.
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PMID:Isolation of collagen from the skins of Ehlers-Danlos syndrome-affected dogs by acetic acid extraction and pepsin digestion. 677 75

The enzymatic degradation of insoluble elastin has been studied at several pH values using purified pepsin and cathepsin D, and neutrophil extracts. Pepsin degraded elastin throughout the pH range of 1.2-4.0 with the optimum pH below 2.0. Molecular sieve chromatography and gel electrophoresis indicated that a spectrum of molecular weight degradation products was produced. The degradation by pepsin was inhibited by sodium dodecyl sulfate (SDS), NaCl and pepstatin. Cathepsin D, which, like pepsin, degrades hemoglobin at acid pH and is inhibited by pepstatin, had no activity against insoluble elastin in the pH range of 3.2-7.2. Extracts of neutrophils degraded elastin above pH 4.0. The pH profile of elastin degradation by neutrophil extracts generally followed that of purified human leukocyte elastase. Our results suggest that during alimentation or pulmonary aspiration of gastric contents, extracellular elastin may be digested by gastric juice at acid pH. Inflammatory cells would not appear to be capable of contributing to such actions until local pH approaches neutrality. Cathepsin D, a major constituent of inflammatory cells, does not digest all types of connective tissue proteins.
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PMID:The enzymatic digestion of elastin at acidic pH. 678 96

Digestion of 18S and 14S acetylcholinesterase from eel electric organ with pepsin at 15 degrees C for 6 h results in extensive degradation of the catalytic subunits, but a major portion of the collagen-like tail structure associated with these enzyme forms resists degradation. The pepsin-resistant structures partially aggregate and can be isolated by gel exclusion chromatography on Sepharose CL-6B in buffered 1 M sodium chloride. The largest structure, denoted F3, has a molecular weight of 72 000 according to gel electrophoresis in sodium dodecyl sulfate and is composed of three 24 000 molecular weight polypeptides linked by intersubunit disulfide bonds. This structure is largely, but not completely, a collagen-like triple helix as indicated by a circular dichroism spectrum typical of triple-helical collagen and an amino acid composition characterized by 27% glycine, 5% hydroxyproline, and 5% hydroxylysine. Continued pepsin action results in degradation of the disulfide linkage region such that disulfide-linked dimers F2 and finally F1 monomers become the predominant forms in sodium dodecyl sulfate. Digested samples in which either F3 or F2 predominate have virtually identical circular dichroic spectra and amino acid compositions and generate similar diffuse 24 000 molecular weight polypeptides following disulfide reduction. Thus the intersubunit disulfide linkages in F3 must occur close to the end(s) of the fragment polypeptide chains. Pepsin conversion of F3 to F2 is particularly accelerated between 25 and 30 degrees C, suggesting that the triple-helical structure in the disulfide linkage region undergoes thermal destabilization in this temperature range. Digestion at 40 degrees C yields presumably triple-helical F1 structures devoid of disulfide linkages, although their degradation to small fragments can be detected at this temperature. The question of whether the three tail subunits that give rise to F1 polypeptides are identical remains open.
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PMID:Characterization of pepsin-resistant collagen-like tail subunit fragments of 18S and 14S acetylcholinesterase from Electrophorus electricus. 678 73

The susceptibility of human type V collagen to several neutral proteases was examined. Thrombin cleaved both the alpha 1(V) and alpha 2(V) chains of this protein at 34 degrees C, producing two pairs of fragments with apparent molecular weights of 95000 and 10000 on sodium dodecyl sulfate--polyacrylamide gel electrophoresis. Two-dimensional 125I-labeled peptide mapping of the larger fragments demonstrated that the upper band [which comigrated with alpha 1(I)] was derived from both the alpha 1(V) and alpha 2(V) chains, while the other component [which comigrated with alpha 2(I) was a product of alpha 1(V) alone. Cleavage of type V collagen, containing alpha 3(V) chains, with thrombin produced an analogous pattern with three high molecular weight bands. Chymotrypsin and trypsin cleaved type V collagen at 37 degrees C but not at lower temperatures. Digestion of type V collagen with elastase at 37 degrees C resulted in selective proteolysis of alpha 2(V), leaving alpha 1(V) essentially intact. Pepsin treatment of type V collagen from which alpha 2(V) had been removed by elastase treatment resulted in nearly complete degradation of alpha 1(V). These data support the hypothesis that a major fraction of native type V collagen is a heteropolymer with the chain composition [alpha 1(V)]2 alpha 2(V). Cleavage of type V collagen by thrombin may have physiologic significance in that breakdown of pericellular matrix may be an important step in the response of a tissue to injury.
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PMID:Susceptibility of type V collagen to neutral proteases: evidence that the major molecular species is a thrombin-sensitive heteropolymer, [alpha 1(V)]2 alpha 2(V). 679 86

The present study was designed to compare collagen synthesized by rabbit lens epithelial cells in culture with rabbit lens capsule collagen. Confluent monolayers of rabbit lens epithelial cells were established. Incorporation of [3H]-proline into glycoproteins secreted into the medium and cell surface components were analyzed in the presence of protease inhibitors. Gel filtration chromatography on sodium dodecyl sulfate--agarose (Bio-gel A-5m) of [3H]-labeled newly synthesized proteins by lens epithelial cells in culture resolved into a single precursor of approximate molecular weight of 160,000 daltons. Neither the medium nor the cell layer showed any evidence of low-molecular weight hydroxyproline-containing material. Limited pepsin digestion of this material cleaved the higher molecular weight chains into smaller components ranging from 25,000 to 110,000 daltons. Pepsin digestion and direct extraction of the collagenous components of the rabbit lens capsule revealed materials of high--molecular weight proteins similar to that synthesized in culture. Low--molecular weight (55,000 daltons) protein was only detected in lens capsules after prolonged pepsin digestion. S-Carboxylation of the lens capsules collagens did not affect their mobilities, but repepsinization gave rise to 110,000 dalton protein, although no significant changes in the amino acid composition were noticed. The absence of synthesis of low--molecular weight protein by cell culture and the presence of low--molecular weight components only after prolonged pepsin digestion of lens capsule could be the result of unusual susceptibility of the basement membrane collagens to pepsin attack.
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PMID:Structure and biosynthesis of rabbit lens capsule collagen. 681 26

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

A collagenous protein could be precipitated by (NH4)2SO4 from the culture medium of a murine teratocarcinoma-derived cell line (Ko, C.Y., Johnson, L.D. and Priest, R.E. (1979) Biochim. Biophys. Acta 581, 252-259). Further purification of this protein was achieved by combining DEAE-cellulose chromatography with either CM-cellulose or molecular sieve chromatography. The collagenous polypeptides had subunit molecular weights of 160 000, if determined by molecular sieve chromatography, or 190 000, if determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and they are not linked by disulphide bridges. Amino acid composition of this collagen is similar to that of a murine type IV collagen isolated from EHS sarcoma (Timpl et al. (1978) Eur. J. Biochem. 84, 43-52). The most prominent peptides resulting from cleavage of the protein by CNBr had estimated molecular weights of 25 000, 23 000, 11 700 and 9400. Pepsin treatment of this collagen under non-denaturing conditions produced three major fragments having molecular weights of 70 000, 45 000 and 43 000. We conclude that the collagen secreted by the murine teratocarcinoma-derived cell culture is a type IV basement membrane collagen. Therefore, this culture system should provide a continuous source of type IV collagen, which may be used to study the interaction of this collagen with other basement membrane components.
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PMID:Purification and characterization of a collagenous protein secreted by a murine teratocarcinoma-derived cell line. 710 99

Pepsin (3.6 nmol) was detected by the following three-step procedure: (i) reaction with a 20-fold molar excess of BO-IMI (a fluorophore containing a reactive imidazole group) in the presence of a 150-fold molar excess of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and 2% sodium dodecyl sulfate; (ii) gel filtration (spin column) to remove most of the residual BO-IMI; and (iii) capillary electrophoresis with laser-induced fluorescence detection. For the latter step, 8.5 x 10(-7) of the original sample was injected. BO-IMI/EDC targets phosphomonoesters and does not label albumin (prior knowledge). Progressive dephosphorylation of pepsin with acid phosphatase reduced its labeling with BO-IMI. Thus, the BO-IMI, as intended, labels the phosphate group on pepsin. Such BO-IMI labeling should be useful in general for studying phosphoproteins and phosphopeptides.
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PMID:Phosphate-specific fluorescence labeling of pepsin by BO-IMI. 750 26

As bovine collagen is currently being scrutinized as to its immunogenicity in clinical use, a human source collagen, human amnion collagen (HAC), has been developed in our laboratory as an injectable biomaterial for soft tissue augmentation. Pepsin-extracted human amnion collagen was highly purified and reconstituted. Gamma irradiation was employed to ensure complete sterility and to produce cross-linking in collagen chains to improve implant persistence without the use of chemical additives. The purity and characteristics of human amnion collagen were proven by amino acid assay, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, immune blotting, and collagenase digestion. Animal studies comparing both irradiated and nonirradiated amnion collagen to bovine collagen (Zyderm and Zyplast) were carried out in a rat model. Humoral immunity was evaluated by examining the sera for antibody reactivity towards the implanted human collagen by the ELISA test. Insignificant antibody levels against human amnion collagen were found. Animal observation revealed fibroplasia, vascular infiltration, and the development of adipocytes with the implant as well as a lack of inflammatory response following up to 12 months of implantation. The persistence rate of our human amnion collagen was equal to, or even longer than, that of both types of bovine collagen implants.
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PMID:Human amnion collagen for soft tissue augmentation--biochemical characterizations and animal observations. 812 34

To investigate the nature of the 140 kDa glycoprotein in the trabecular meshwork, polypeptides were extracted with either urea/sodium dodecyl sulfate (SDS)/beta-2-mercaptoethanol (BME) or guanidine hydrochloride followed by pepsin digestion. After electrophoresis and immunoblotting with anti-type-VI-collagen antibodies, a single fraction of molecular weight 140 kDa was identified in the urea/SDS/BME extracts. Pepsin solubilization revealed two immunoreactive fractions (molecular weights 75 and 85 kDa) that comigrated with purified, pepsin-solubilized type VI collagen. By using the polymerase chain reaction (PCR) and primers specific for the alpha 2(VI) chain of type VI collagen, a single PCR product was obtained, which corresponded to the expected size of 137 base pairs, from the total RNA extracted from the trabecular meshwork ex vivo. Southern hybridization with the antisense oligonucleotide probe of the alpha 2(VI) chain confirmed that the amplified sequence was specific. The results show that the trabecular meshwork contains a significant amount of type VI collagen and that trabecular cells express the mRNA coding for the alpha 2(VI) chain of this glycoprotein. The presence of type VI collagen in the trabecular meshwork is implicated in cell-extracellular matrix interactions at this site, and its abnormal accumulation in glaucomatous and aging eyes probably signifies a defect in the function of the trabecular cells in these eyes.
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PMID:Identification of type VI collagen in the trabecular meshwork and expression of its mRNA by trabecular cells. 815 10


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