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

Inside APCs, MHC class II molecules associate with antigenic peptides before reaching the cell surface. This association takes place in compartments of the endocytic pathway, more related to endosomes or lysosomes depending on the cell type. Here, we compared MHC class II transport from endosomal vs lysosomal compartments to the plasma membrane. We show that transport of MHC class II molecules to the cell surface does not depend on the cytosolic domains of the alpha- and beta-chains. In contrast, the stability of the alphabeta-peptide complexes determined the efficiency of transport to the cell surface from lysosomal, but not from endosomal, compartments. In murine B lymphoma cells, SDS-unstable and -stable complexes were transported to the cell surface at almost similar rates, whereas after lysosomal relocalization or in a cell line in which MHC class II molecules normally accumulate in lysosomal compartments, stable complexes were preferentially addressed to the cell surface. Our results suggest that when peptide loading occurs in lysosomal compartments, selective retention and lysosomal degradation of unstable dimers result in the expression of highly stable MHC class II-peptide complexes at the APC surface.
 ...
PMID:MHC class II transport from lysosomal compartments to the cell surface is determined by stable peptide binding, but not by the cytosolic domains of the alpha- and beta-chains. 972 1

Sarcosine reductase is the only reductase system present in Tissierella creatinophila when grown on creatinine plus formate. The acetyl-phosphate-forming component protein C was purified to homogeneity. SDS-PAGE of the purified protein revealed two protein bands with apparent mol. masses of 62 and 50 kDa. The N-terminal amino acid sequence of the two subunits was determined. Antibodies raised against each of the subunits of protein C from Eubacterium acidaminophilum cross-reacted with the corresponding protein present in T. creatinophila, Clostridium litorale and Clostridium sporogenes. The arsenate-dependent hydrolysis of acetyl phosphate catalyzed by protein C was partly inhibited by antibodies directed against the large subunit. Antibodies raised against the small subunit were twice as effective, which indicates that this subunit is the primary site of acetyl transfer from acetyl phosphate. The protein A component of the sarcosine reductase of T. creatinophila was purified to homogeneity by cochromatography with thioredoxin reductase on DEAE-Sephacel, hydroxylapatite, Q-Sepharose, and Sephacryl 100-HR. Protein A had an apparent mol. mass of 21 kDa. Its N-terminal amino acid sequence showed high similarities to that of other proteins A. Initial steps for the purification and preliminary characterization of the sarcosine-specific, substrate-binding protein Bsarcosine component of T. creatinophila indicated the involvement of a 50-kDa protein.
 ...
PMID:Sarcosine reductase of Tissierella creatinophila: purification and characterization of its components. 979 88

The effect of prostaglandins (PGA1 and PGB2) on the replication of Mayaro virus was studied in Vero cells. PGA1 and PGB2 antiviral activity was found to be dose-dependent. However, while 10 micrograms/ml PGB2 inhibited virus yield by 60%, at the same dose PGA1 suppressed virus replication by more than 90%. SDS-PAGE analysis of [35S]-methionine-labelled proteins showed that PGA1 did not alter cellular protein synthesis. In infected cells, PGA1 slightly inhibited the synthesis of protein C, while drastically inhibiting the synthesis of glycoproteins E1 and E2.
 ...
PMID:Inhibition of Mayaro virus replication by prostaglandin A1 and B2 in Vero cells. 987 77

Factor VIIa, in complex with tissue factor (TF), is the serine protease responsible for initiating the clotting cascade. This enzyme complex (TF/VIIa) has extremely restricted substrate specificity, recognizing only three previously known macromolecular substrates (serine protease zymogens, factors VII, IX, and X). In this study, we found that TF/VIIa was able to cleave multiple peptide bonds in the coagulation cofactor, factor V. SDS-PAGE analysis and sequencing indicated the factor V was cleaved at Arg679, Arg709, Arg1018, and Arg1192, resulting in a molecule with a truncated heavy chain and an extended light chain. This product (FVTF/VIIa) had essentially unchanged activity in clotting assays when compared to the starting material. TF reconstituted into phosphatidylcholine vesicles was ineffective as a cofactor for the factor VIIa cleavage of factor V. However, incorporation of phosphatidylethanolamine in the vesicles had little effect over the presence of 20% phosphatidylserine. FVTF/VIIa was as sensitive to inactivation by activated protein C (APC) as thrombin activated factor V as measured in clotting assays or by the appearance of the expected heavy chain cleavage products. The FVTF/VIIa could be further cleaved by thrombin to release the normal light chain, albeit at a significantly slower rate than native factor V, to yield a fully functional product. These studies thus reveal an additional substrate for the TF/VIIa complex. They also indicate a new potential regulatory pathway of the coagulation cascade, i.e., the production of a form of factor V that can be destroyed by APC without the requirement for full activation of the cofactor precursor.
 ...
PMID:Factor VIIa/tissue factor generates a form of factor V with unchanged specific activity, resistance to activation by thrombin, and increased sensitivity to activated protein C. 1002 63

Increased levels of the acute phase protein C-reactive protein (CRP) in plasma may indicate severe acute abdominal disease, risk of serious postoperative complications or malignancy; serial measurements may indicate postoperative complications, relapse of intra-abdominal sepsis and complications during acute pancreatitis. The increase in CRP is an unspecific acute phase reaction, however, and low levels do not exclude these conditions. These facts are important obstacles to the clinical routine use of CRP measurements. The aim of this study was to look for possible biochemical microheterogeneity of CRP in single plasma samples from various large groups of patients to overcome these problems. Two-hundred-and-twelve patients with acute abdominal diseases, 274 patients with various forms and stages of cancer and 134 patients operated on due to benign diseases, were studied. The biochemical studies included SDS-PAGE, native PAGE and gel filtration for molecular weight determinations, isoelectric focusing and crossed immuno-electrophoresis for electrophoretic mobility studies and Concavalin A and ACA 34 as intermediary gels for possible lectin binding or complexation. Western blot analysis was also used to identify CRP. In summary, however, these more elaborate biochemical methods could not disclose any microheterogneity of CRP in plasma and thus did not add any diagnostic information to the crude levels.
 ...
PMID:No microheterogenous changes of plasma C-reactive protein found in man during various diseases. 1046 68

We have selected the Streptoalloteichus hindustanus bleomycin-resistance protein ShBle, a 28-kDa homodimer, as a scaffold for the display of bioactive peptides and other peptide epitopes. To create a monomeric scaffold, we investigated the effect of mutating residue proline 9 to glycine. This residue plays a critical role in ShBle dimerization by affecting the position of the eight N-terminal residues which secure the interaction between the monomeric subunits. We demonstrate that this mutation weakens the dimerization interaction, resulting in establishment of a stable equilibrium between monomeric and dimeric ShBle species in solution. Circular dichroism and SDS-PAGE data indicate that the Pro9Gly mutation does not disrupt the structure of the molecule. Production of a fully monomeric form of ShBle required complete removal of the eight-residue N-terminal peptide, and the interaction across the now solvent-exposed hydrophobic interface of the ShBle monomer was insufficient to drive dimerization. To demonstrate efficient display of epitope tags on the ShBle protein, we displayed dual-octapeptide FLAG tags at the protein C-terminus. These additions did not interfere with protein folding or activity. The resulting ShBle scaffold was used to compare the efficiency of two commercial FLAG-specific antibodies by biosensor.
 ...
PMID:Utilization of the Streptoalloteichus hindustanus resistance determinant ShBle as a protein framework: effect of mutation upon ShBle dimerization and interaction of C-terminal displayed peptide epitopes. 1083 17

The zinc endopeptidases mutalysin I (100 kDa) and mutalysin II (22.5 kDa) have been previously isolated from bushmaster (Lachesis muta muta) snake venom. Hemorrhagic activity was observed with as little as 0.5 microg (2000 units/mg) and 17.8 microg (56.2 units/mg) for mutalysin I and II, respectively. Additionally, the proteases hydrolyse the Aalpha>Bbeta chain of fibrinogen without clot formation. The specific fibrinogenolytic activity was estimated as 5. 25 and 16.3 micromol fibrinogen/min/micromol protein for mutalysin I and II, respectively. In vitro, the enzymes act directly on fibrin and are not inhibited by serine proteinase inhibitors (SERPINS). Analysis by SDS-PAGE of fibrin hydrolysis by both enzymes showed that mutalysin II (0.22 microM) completely digested the alpha- and gamma-gamma chains and partially the beta-chain (in 120 min incubation). In contrast, mutalysin I (three fold higher concentration than mutalysin II) hydrolyzed selectively the alpha-chain of fibrin leaving the beta and gamma-gamma chains unaffected. Unlike with the plasminogen activator-based thrombolytic agents (e.g., streptokinase), mutalysins do not activate plasminogen. Neither enzyme had an effect on protein C activation. Mutalysin II does not inhibit platelet aggregation in human PRP induced by collagen or ADP. However, mutalysin I showed a selective inhibitory effect on collagen-induced aggregation of human PRP; it did not affect platelet aggregation with ADP as the agonist. The present investigation demonstrates that both native and EDTA-inactivated mutalysin I dose dependently blocked aggregation of human PRP elicited by 10 microg/mL of collagen with an IC(50) of 180 and 580 nM, respectively. These studies suggest that, in addition to the metalloprotease region of mutalysin I, the disintegrin-like domain also participates in the inhibitory effect. The proteolytic activity of mutalysin II against dimethylcasein and fibrin was completely abolished by alpha2-macroglobulin (alpha2-M). The stoichiometry of inhibition was 1.0 mol of enzyme per mol of alpha2-M. In contrast, the proteolytic effect of mutalysin I against the same substrates was not significantly inhibited by alpha2-M. Therefore, the data explain why mutalysin I contributes significantly not only to local but also to systemic bleeding associated with the observed pathological effects of the venom.
 ...
PMID:Action of metalloproteinases mutalysin I and II on several components of the hemostatic and fibrinolytic systems. 1096 87

To characterize the putative biochemical modifications induced by the Ser 460 to Pro (Heerlen) mutation in protein S (PS), we expressed both wild-type (wt) and mutated recombinant PS in HEK cells. In SDS-polyacrylamide gels, r-PS Heerlen migrated at 71 kDa whereas r-wt PS migrated at 73 kDa, a difference abolished after deglycosylation by N-glycosidase, suggesting that the Ser 460 Pro mutation abolishes N-glycosylation of Asn 458. The affinity of r-wt PS and r-PS Heerlen for C4b-binding protein (C4b-BP) and for phospholipid vesicles was similar. Neither the enhancement of APC-dependent prolongation of the APTT, nor the specific enhancement of FVa and FVIIIa proteolysis by APC in purified systems was affected by the mutation. However, the Ser 460 Pro mutation induced a slight conformational change in the SHBG domain of the PS molecule, as shown by reduced binding affinity for monoclonal antibodies. The type III phenotype associated with the Heerlen mutation might thus result from a slightly modified rate of synthesis or catabolism. The resulting moderate decrease in the circulating PS concentration may modify the equilibrium between free PS and C4b-BP/PS complexes.
 ...
PMID:Expression and characterization of recombinant protein S with the Ser 460 Pro mutation. 1105 20

Recombinant factor VIII SQ (r-VIII SQ), ReFacto, is a recombinant factor VIII product similar to the smallest active factor VIII protein found in plasma-derived factor VIII (p-VIII) concentrates. The protein comprises two polypeptide chains of 80 and 90 kDa and lacks the major part of the heavily glycosylated B-domain i.e. amino acids Gln744 to Ser1637. r-VIII SQ retains six potential glycosylation sites for N-linked oligosaccharides at asparagine residues 41, 239, 582, 1685, 1810 and 2118. We describe a thorough comparison of the characteristics of r-VIII SQ with those of p-VIII. The primary and secondary structures of r-VIII SQ were in good agreement with that of B-domain-deleted p-VIII (p-VIII-LMW) as shown by SDS-PAGE, Western blotting with antifactor VIII antibodies, tryptic mapping, amino acid sequence analysis and circular dichroism spectroscopy. A few divergences also existed. Thus r-VIII SQ was shown to contain a small amount of the single chain primary translation product of 170 kDa and also the product specific sequence of 14 amino acids, the SQ-link, in the C-terminal end of the 90 kDa chain. It was shown that r-VIII SQ had a high specific activity of about 14,000 IU VIII:C/mg as determined by use of a chromogenic substrate assay. The r-VIII SQ protein was comparable to p-VIII forms with a retained B-domain, in terms of potency measured by a chromogenic substrate or a two-stage clotting assay, in interactions with thrombin, and with activated protein C (APC) in combination with Protein S. The ability of r-VIII SQ to participate as a cofactor in factor Xa generation in a mixture of factors IXa and X, phospholipid and calcium was in conformity with that of p-VIII. Furthermore r-VIII SQ had a good binding capacity for phospholipid vesicles and von Willebrand factor (vWF) as shown in gel filtration studies. The same kinetics in binding to von Willebrand factor was found for r-VIII SQ and p-VIII as determined by real-time biospecific interaction analysis (BIA) with use of the BIAcore instrument. The apparent association rate constant was 4 x 10(6) M(-1)s(-1). Two dissociation rate constants were found, 1 X 10(-2)s(-1) and 4 x 10(-4)s(-1). The results extend the present knowledge that the factor VIII B-domain is dispensable for the factor VIII cofactor function in hemostasis.
 ...
PMID:Structural and functional characteristics of the B-domain-deleted recombinant factor VIII protein, r-VIII SQ. 1120 95

Factors VII, IX, and X play key roles in blood coagulation. Each protein contains an N-terminal gamma-carboxyglutamic acid domain, followed by EGF1 and EGF2 domains, and the C-terminal serine protease domain. Protein C has similar domain structure and functions as an anticoagulant. During physiologic clotting, the factor VIIa-tissue factor (FVIIa*TF) complex activates both factor IX (FIX) and factor X (FX). FVIIa represents the enzyme, and TF represents the membrane-bound cofactor for this reaction. The substrates FIX and FX may utilize multiple domains in binding to the FVIIa*TF complex. To investigate the role of the EGF1 domain in this context, we expressed wild type FIX (FIX(WT)), FIX(Q50P), FIX(PCEGF1) (EGF1 domain replaced with that of protein C), FIX(DeltaEGF1) (EGF1 domain deleted), FX(WT), and FX(PCEGF1). Complexes of FVIIa with TF as well as with soluble TF (sTF) lacking the transmembrane region were prepared, and activations of WT and mutant proteins were monitored by SDS-PAGE and by enzyme assays. FVIIa*TF or FVIIa*sTF activated each mutant significantly more slowly than the FIX(WT) or FX(WT). Importantly, in ligand blot assays, FIX(WT) and FX(WT) bound to sTF, whereas mutants did not; however, all mutants and WT proteins bound to FVIIa. Further experiments revealed that the affinity of the mutants for sTF was reduced 3-10-fold and that the synthetic EGF1 domain (of FIX) inhibited FIX binding to sTF with K(i) of approximately 60 microm. Notably, each FIXa or FXa mutant activated FVII and bound to antithrombin, normally indicating correct folding of each protein. In additional experiments, FIXa with or without FVIIIa activated FX(WT) and FX(PCEGF1) normally, which is interpreted to mean that the EGF1 domain of FX does not play a significant role in its interaction with FVIIIa. Cumulatively, our data reveal that substrates FIX and FX in addition to interacting with FVIIa (enzyme) interact with TF (cofactor) using, in part, the EGF1 domain.
 ...
PMID:The N-terminal epidermal growth factor-like domain in factor IX and factor X represents an important recognition motif for binding to tissue factor. 1172 40


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>