Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0272170 (SDS)
 50,377 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The proteins and polysaccharides which are left adherent to the tissue culture substrate after EGTA-mediated removal of normal, virus-transformed, and revertant mouse cells (so-called SAM, or substrate-attached material), and which have been implicated in the cell-substrate adhesion process, have been characterized by SDS-PAGE and other types of analyses under various conditions of cell growth and attachment. The following components have been identified in SAM: 3 size classes of hyaluronate proteoglycans; glycoprotein Co (the LETS glycoprotein); protein Ca (a myosin-like protein); protein Cb(MW 85,000); protein C1 (MW 56,000, which is apparently not tubulin); protein C2 (actin); proteins C3-C5 (histones) which are artifactually bound to the substrate as a result of EGTA-mediated leaching from the cell; and proteins Cc, Cd, Ce, and Cf. The LETS glycoprotein (Co) and Cd appear in newly-synthesized SAM (which is probably enriched in "footpad" material--"footpads" being focal areas of subsurface membraneous contact with the substrate in greater relative quantities than in the SAM accumulated over a long period of time (which is probably enriched in "footprint" material--remnants of footpads left behind as cells move across the substrate). CO and Cd turn over very rapidly following short radiolabeling periods during chase analysis. The SAM's deposited during a wide variety of cellular attachment and growth conditions contained the same components in similar relative proportions. This may indicate well-controlled and coordinate deposition of a cell "surface" complex involving the hyaluronate proteoglycans, the LETS glycoprotein, actin-containing microfilaments with associated proteins, and a limited number of additional proteins in the substrate adhesion site. Evidence indicates that SAM is the remnant of "footpad" vesicles by which the cell adheres to the substrate and that EGTA treatment weakens the subsurface cytoskeleton, allowing these footpad vesicles to be pinched off from the rest of the cell. Three different models of cell-substrate adhesion are presented and discussed.
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PMID:Molecular composition and origin of substrate-attached material from normal and virus-transformed cells. 100 70

Modification of protein Ag by proteolysis is one of the principal steps in the presentation of Ag to Th cells. However, little is known about the enzymes participating in these events, their specificity or the characteristics of the natural fragments that they produce. Cathepsin D (CD) is an aspartyl protease identified in endosomes of APC. In this report, the role of CD in the processing of OVA has been investigated. OVA digested in vitro with purified CD was able to stimulate IL-2 secretion by three different OVA-specific I-Ad restricted Th cell hybridomas when it was presented by fixed APC. The digest of OVA was recognized in the context of I-Ad, but not by I-Ak-restricted OVA-specific Th cells. No difference was observed in the ability of OVA digested with CD to stimulate Th cells in the absence of FCS or in the presence of protease inhibitors indicating that extracellular proteases were not likely to contribute to processing of OVA. Taken together, these results suggest that CD is necessary and sufficient for the generation of an antigenic epitope from OVA. A fragment containing the epitope was isolated from the OVA digest by reverse phase HPLC. This fragment, which migrates in SDS-PAGE as a 10-kDa polypeptide, is a potent epitope. Its capacity to activate Th cells is compared to that of the tryptic peptide OVA323-339.
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PMID:Role of cathepsin D in antigen presentation of ovalbumin. 132 88

Cathepsin G was used in vitro to digest human factor VII and factor IX. Clotting assays indicated that the proteinase affected a rapid loss in coagulant activity while in the presence of calcium ions the activity was almost totally protected. SDS-polyacrylamide gel electrophoresis indicated the removal of a peptide from each zymogen, VII-L from factor VII and IX-L from factor IX. This lead to the formation of VII-H and IX-H respectively. N-terminal analysis of the VII-H and IX-H products and COOH-terminal analysis of the VII-L and IX-L products confirmed that cathepsin G had cleaved position Phe40:Trp41 in factor VII and factor IX. The cleavage site is the same as that when cathepsin G is reacted with factor II, factor X and protein C. The unique action of cathepsin G may be part of a regulatory system for controlling the coagulant activity of vitamin K dependent clotting in vivo.
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PMID:Cathepsin G, a regulator of human vitamin K, dependent clotting factors and inhibitors. 144 May 18

Rat liver macrophages express a galactose-specific receptor which mediates endocytosis of particles or neuraminidase-treated blood cells. From rat serum we now have isolated a galactose-specific lectin by affinity chromatography. Comparative analysis of this serum galactose-binding protein with the galactose-specific particle receptor protein purified from rat liver macrophages and with the acute-phase protein C-reactive protein (CRP) revealed a close relation or identity of these proteins. An apparent molecular weight of 30 kilodaltons was determined for all three proteins by SDS-PAGE under reducing conditions and of about 130 kilodaltons by native PAGE. All three proteins exhibit the same pentameric, ring-shaped structure. Antibodies raised against the serum galactose-binding protein or against the macrophage receptor did cross-react. Monoclonal antibodies raised against rat CRP labeled liver macrophage but not hepatocyte surfaces and reacted with all three isolated proteins in a Western blot assay. Furthermore, the galactose-specific particle receptor could be functionally replaced by purified CRP. Northern blot analysis showed that the CRP is not synthesized in the macrophages but appears to be acquired from hepatocytes or blood. We now conclude that a membrane-bound form of CRP functions as the recycling galactose-specific particle receptor in rat liver Kupffer cells.
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PMID:A membrane-bound form of the acute-phase protein C-reactive protein is the galactose-specific particle receptor on rat liver macrophages. 165 73

Thrombin, the final enzyme of the coagulation system, also influences profibrinolytic activity by several mechanisms. These include cellular release of tissue plasminogen activator, activated protein C-induced fibrinolysis, and inactivation of plasminogen activator inhibitor, type 1 (PAI-1). In this report, the role of thrombin in the regulation of PAI-1 is investigated. Our studies demonstrate that thrombin inactivation of PAI-1 occurs via an enzymatic mechanism rather than an enzyme-inhibitor complex mechanism. Evidence to support this conclusion is: (1) concomitant analysis of PAI-1 and thrombin activities demonstrate decreased PAI-1 activity but no loss of thrombin activity; (2) no visible thrombin--PAI-1 complexes by SDS-PAGE analysis; and (3) lack of formation of 125I-thrombin-PAI-1 complexes. Thrombomodulin, a thrombin binding cofactor that modifies thrombin's functions, did not influence the inactivation of PAI-1 by thrombin. We propose that thrombin enzymatically inactivates PAI-1 without forming a stable enzyme-inhibitor complex. The reaction is not affected by thrombomodulin. Overall this reaction occurs so slowly that it is not physiologically relevant without some modifying factor(s).
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PMID:Thrombin and the thrombin-thrombomodulin complex interaction with plasminogen activator inhibitor type-1. 165 27

A low molecular weight platelet inhibitor of factor XIa (PIXI) has been purified 250-fold from releasates of washed and stimulated human platelets. Molecular weight estimates of 8400 and 8500 were determined by gel filtration and SDS-polyacrylamide gel electrophoresis, respectively, although a second band of Mr 5000 was present upon electrophoresis. The inhibitor does not appear to be one of the platelet-specific, heparin-binding proteins, since it neither bound to nor was affected by heparin. An amount of PIXI which inhibited by 50% factor XIa cleavage of the chromogenic substrate S2366 (Pyr-Glu-Pro-Arg-pNA-2H2O) only slightly inhibited (5-9%) factor XIIa, plasma kallikrein, plasmin, and activated protein C and did not inhibit factor Xa, thrombin, tPA, or trypsin, suggesting specificity for factor XIa. Kinetic analyses of the effect of PIXI on factor XIa activity demonstrated mixed-type, noncompetitive inhibition of S2366 cleavage and of factor IX activation with Ki's of 7 x 10(-8) and 3.8 x 10(-9) M, respectively. Immunoblot analysis showed that PIXI is not the inhibitory domain of protease nexin II, a potent inhibitor of factor XIa also secreted from platelets. Amino acid analysis showed that PIXI has no cysteine residues and, therefore, is not a Kunitz-type inhibitor. PIXI can prevent stable complex formation between alpha 1-protease inhibitor and factor XIa light chain as demonstrated by SDS-polyacrylamide gel electrophoresis. The inhibition by PIXI of factor XIa-catalyzed activation of factor IX and its capacity to prevent factor XIa inactivation by alpha 1-protease inhibitor, combined with the specificity of PIXI for factor XIa among serine proteases found in blood, suggest a role for PIXI in the regulation of intrinsic coagulation.
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PMID:A low molecular weight platelet inhibitor of factor XIa: purification, characterization, and possible role in blood coagulation. 173 24

Human polymorphonuclear leucocyte cathepsin G was used in vitro to digest human protein C. Anticoagulant assays showed that the proteinase affected a rapid loss in the anticoagulant activity of the zymogen. Calcium ions almost totally protected the anticoagulant activity of protein C against the action of cathepsin G. Polyacrylamide gel electrophoresis, in the presence of SDS (non reducing conditions), indicated that the proteolytic action of cathepsin G lead, to the removal of a peptide of low molecular mass (PC-L'') from protein C with the consequent formation of a product of high molecular mass (PC-P). Electrophoresis in the presence of SDS (reducing conditions) showed that the PC-L'' fragment was removed from the light chain of protein C (PC-L). N-terminal amino acid analysis of the PC-P product and COOH terminal amino acid analysis of PC-L'' product confirmed that cathepsin G had cleaved position Phe40:Trp41 in the light chain. This cleavage was responsible for removal of the gamma-carboxyglutamic acid (gla) containing region of protein C.
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PMID:Cathepsin G can produce a gla-domainless protein C in vitro. 175 93

Three strains of spirochetes (IKA1 to 3) were isolated from the midgut of Ixodes ovatus collected in the Ikawa region of the northern part of Shizuoka, Japan. These isolates had eight flagella, and their size and other morphological features were similar to Borrelia burgdorferi. They showed similar motility and reacted with monoclonal antibody (MAb) H9724 against borrelial flagella and with MAb H5332 against the outer surface protein A. These strains showed similar SDS-PAGE profiles to that of B. burgdorferi strain B31 and P/Bi isolated in the U.S.A. and Europe, respectively. Immunoblot with Lyme disease patient serum showed positive reactions with the flagella (41 Kilodalton, kDa), protein C (20 to 22 kDa), and outer surface protein A (29 kDa) of the isolates. Immunological properties, morphological characteristics, and epidemiological features revealed that these isolates were B. burgdorferi.
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PMID:Immunological properties of Borrelia burgdorferi isolated from the Ixodes ovatus in Shizuoka, Japan. 177 92

A recombinant Factor VIII (Factor VIII-delta II) consists of a unique polypeptide chain of 165 kDa deleted from the major part of the B-domain and from the cleavage site at Arg-1648-Glu-1649 found in plasma-derived Factor VIII. It was expressed in mammalian cells in serum-free medium containing von Willebrand factor and purified by a one-step immunopurification. The recombinant Factor VIII was characterized as a single active peak when subjected to f.p.l.c., in contrast with the plasma-derived molecule. Its coagulant activity was decreased in the presence of EDTA, suggesting that a bivalent ion is required, as for plasma-derived Factor VIII. The activation by thrombin and the inactivation by activated protein C were studied and the resulting molecular forms were analysed by f.p.l.c. and SDS/PAGE. The results clearly demonstrate that, despite the structural differences between plasma-derived and recombinant Factor VIII, activation and inactivation of Factor VIII-delta II generate proteolysed complexes similar to that described for plasma-derived Factor VIII. Thus this deleted recombinant Factor VIII, which is processed similarly to plasma-derived Factor VIII, should be normally integrated in the regulation system of Factor X activation in the blood-coagulation cascade.
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PMID:Structural and functional characterization of Factor VIII-delta II, a new recombinant Factor VIII lacking most of the B-domain. 190 11

Human anticoagulant vitamin-K-dependent protein S was expressed in mouse C127 cells using a bovine papilloma virus vector system. A full-length cDNA construct was introduced into the vector in the 5' untranslated region of the mouse metallothionein-I gene. Transfected cells expressed approximately 10 micrograms/ml of the recombinant protein which was purified by ion-exchange chromatography followed by affinity chromatography using Ca2(+)-dependent monoclonal antibodies against the region of protein S containing 4-carboxyglutamic acid. Recombinant protein S was structurally and functionally similar to protein S purified from plasma. On SDS/polyacrylamide-gel electrophoresis recombinant protein S had a slightly higher molecular mass than plasma protein S. After treatment with endoglycosidase F, the proteins comigrated suggesting the observed molecular mass difference to be due to alterations in the N-linked carbohydrate side chains. Recombinant and plasma protein S demonstrated identical amino-terminal sequences, similar amino acid composition and number of 4-carboxyglutamyl and 3-hydroxyaspartyl/asparaginyl residues. Recombinant protein S had the same affinity for Ca2+ as protein S from plasma and the two proteins had the same activated protein C cofactor activity in a functional assay. In addition, both forms of protein S formed complexes with C4b-binding protein with the same apparent Kd. Protein S is the most extensively post-translationally modified vitamin-K-dependent protein, and all the modifications were carried out in the recombinant DNA system yielding a recombinant protein S with full biological activity.
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PMID:Expression of completely gamma-carboxylated and beta-hydroxylated recombinant human vitamin-K-dependent protein S with full biological activity. 213 11


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