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 molecular interaction of bovine kininogen and its derivatives with papain was investigated. High-molecular-weight kininogen (HMWK) or low-molecular-weight kininogen (LMWK) and inactive papain treated with N-[N-(L-3-trans-carboxyoxiran-2-carbonyl)-L-leucyl]agmatine (E-64) formed, respectively, a complex, which was dissociable on sodium dodecyl sulfate polyacrylamide-gel electrophoresis (SDS-PAGE). The densitometric determination of the bands separated on SDS-PAGE and amino acid analysis of the samples extracted from the electrophoresis gel revealed that the complex between kininogen and papain is formed in a molar ratio of one to one. Moreover, analysis of the inhibition of the caseinolytic activity of papain by these kininogens indicated that HMWK, LMWK, and kinin-free derivatives obtained from both kininogens inhibit active papain with a stoichiometry of 1:1. On the other hand, the papain activity was inhibited by two kinds of cyanogen bromide fragments isolated from the heavy chain of HMWK. These two fragments with Ki values of 38 and 0.64 nM corresponded, respectively, to residue Nos. 47 to 243 and Nos. 244-360 of the HMWK heavy chain. These results suggest that in the intact HMWK and LMWK, one of the two potential reactive sites interacts with papain to form a complex and that the other reactive site becomes active only after separation of the two sites.
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PMID:Molecular interaction of bovine kininogen and its derivatives with papain. 318 63

Dictyostelium myosin was associated into dimers and small oligomers at very low ionic strength, filamentous at intermediate ionic strength, and monomeric in solution conditions of high ionic strength. These different associations were probed by fragmenting myosin with chymotrypsin, trypsin, or V-8 protease. All three proteases digested monomeric myosin giving rise to multiple fragments with a wide range of molecular weights. Filamentous myosin was not digested by the V-8 protease, was preferentially cleaved at a single site in the middle of the heavy chain by chymotrypsin, and was cleaved at several sites by trypsin. If the reaction was carried out in very low ionic strength, however, two of these proteases generated stable fragments of high molecular weight. Electron microscopic analysis of these stable fragments showed that tails were shorter than in intact myosin, indicating that the cleavage sites were in the rod portion of the molecule. Under the same conditions of enzymatic digestion, myosin that had been radio labeled in vivo with 32P was analyzed by SDS-PAGE and autoradiography. By comparing the state of phosphorylation and the size of the stable fragments, it was determined that the heavy chain phosphorylation site was located between 55 and 70 kD from the tip of the myosin tail, near a region where the tail displayed sharp bends.
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PMID:Proteolytic fragmentation of Dictyostelium myosin and localization of the in vivo heavy chain phosphorylation site. 320 94

Unreduced human immunoglobulin G (IgG) which was not aggregated showed anomalous apparent molecular masses on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). It migrated mainly as three distinct bands with apparent molecular masses from 190 to 240 kDa on gels containing 8% polyacrylamide, when denatured at 37 degrees C. Generation of this banding pattern has two reasons: (a) the pattern is a superposition of bands originating from the four IgG subclasses that differ in molecular masses and structures; and (b) the complexity of the band pattern is further increased, because IgG myeloma proteins of the IgG1 and IgG2 subclass migrated as doublets, while IgG3 and IgG4 formed primarily one band with slightly different apparent molecular masses. These properties were independent of the type of light chain in all myeloma proteins studied. Generation of doublets suggests heterogeneities of monoclonal proteins. The two separable protein populations from IgG1 differ in their susceptibility to reduction. Reduction at 37 degrees C cleaved the larger into heavy and light chain, while it generated heavy chain dimer and light chain from the smaller species. Hence, it is possible that monoclonal IgG1 are comprised of at least two subpopulations of molecules with different S-S bonds. Doublet formation of IgG2 remains unexplained, since both species were equally sensitive to reduction. Knowledge on the anomalous properties of IgG on SDS-PAGE is a prerequisite to run immunoblots from unreduced cellular antigens without confounding cell-associated IgG with cellular antigens.
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PMID:Electrophoretic properties of human IgG and its subclasses on sodium dodecyl-sulfate-polyacrylamide gel electrophoresis and immunoblots. 323 61

We reported in the preceding paper [Muno, D., et al. (1987) J. Biochem. 101, 661-669] that the dinitrophenyl group exclusively introduced to SH1 on the 20-kDa fragment of myosin subfragment 1 was cross-linked to the 50-kDa fragment by irradiation, and that limited trypsinolysis of the cross-linked S1 generated an 83-kDa peptide, a cross-linking product between the 20- and 50-kDa fragments. This paper will deal with the location of the cross-linked residue on the 50-kDa fragment. When the 83-kDa fragment labeled at SH2 with a fluorogenic SH reagent was subjected to bromocyanolysis, a main fluorescent band, which implied a cross-linked peptide, appeared in the position with an apparent molecular mass of 18.5-kDa on SDS-PAGE. On the other hand, another cross-linked peptide was obtained from a complete tryptic digest of a 83-kDa fragment rich fraction. Amino acid sequence analysis of the two cross-linked peptides revealed that the DNP moiety attached at SH1 was cross-linked with a residue in the segment of the heavy chain spanning the 485-493 region from the N-terminus of the heavy chain.
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PMID:Photocross-linking from dinitrophenylated SH1 in myosin head. II. Cross-linked site on 50-kDa fragment. 324 Sep 84

Monoclonal antibody 5E2 identifies a new rabbit thymocyte specific cell surface molecule designated R-Ta. SDS-PAGE of molecules immunoprecipitated by 5E2 shows that R-Ta exists as a non-covalently associated hetero-dimer consisting of a light polypeptide chain (mol. wt approximately 12,000) and a bi-molecular species of a heavy chain (mol. wts of 45,000 and 40,000). The difference between the two forms of heavy chain can be attributed to different degrees of glycosylation. Each form of the R-Ta heavy chain has a polypeptide mol. wt of 34,000. At least three N-linked oligosaccharides and no significant O-linked sugars were found associated with R-Ta. Two dimensional electrophoresis of V8 protease peptide maps also indicate that the two forms of the heavy chains are similar, if not identical, in polypeptide primary structure. The light polypeptide was found to be serologically and structurally identical to beta-2-microglobulin. This was demonstrated in a previous study by reaction with goat anti-beta-2-microglobulin antisera. In this investigation the structural identity with beta-2-microglobulin was demonstrated by partial amino terminal sequence analysis. The partial amino acid sequence for 18 steps of the R-Ta heavy chain was also determined. A comparison of the amino acid sequence with other known sequences for the conventional Class I molecules of man, mouse and rabbit did not reveal any homology. Thus R-Ta is a new T-cell surface protein, and like human CD1, carries the unique distinction of thymocyte specificity, is beta-2-microglobulin associated, but is not Class I related.
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PMID:Biochemical properties of a novel rabbit thymocyte specific class I-like antigen. 326 85

We have described previously the IgE-mediated release of kininogenase activity from purified human lung mast cells. Using supernatant fractions from mast cells stimulated with anti-IgE in the presence of deuterium oxide, we have purified this kininogenase to homogeneity by gel filtration and heparin-agarose chromatography and have demonstrated that it is identical to tryptase, the major neutral protease of human lung mast cells. Thus, tryptase and kininogenase activities co-chromatographed through both purification steps with equivalent yields. The final purified kininogenase was free of detectable chymotryptic and carboxypeptidase activities and was identified as tryptase on the basis of sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), amino acid composition and inhibition profile. Three such preparations of tryptase were all capable of releasing kinin from each of two different preparations of purified, single-chain, human low molecular weight kininogen. Interestingly, kinin generation was optimal at pH 5.5 and was enhanced by heparin, which has been reported to stabilize tryptase. SDS-PAGE analysis of kininogen hydrolysis by tryptase revealed the formation of a diffusely stained region in the molecular weight range of 60,000-65,000, rather than a discrete heavy chain band. Under optimal conditions, the three tryptase preparations released 10-12 micrograms kinin/hr/mg but released only 2 micrograms kinin/hr/mg at pH 7.2. HPLC analysis revealed that the kinin released was bradykinin. We conclude that the kininogenase activity from human lung mast cells is attributable to tryptase. The unique pH optimum of this reaction of a serine protease, however, raises doubts as to the physiologic significance of this activity.
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PMID:Identification of human lung mast cell kininogenase as tryptase and relevance of tryptase kininogenase activity. 328 17

Monoclonal antibodies directed to MHC class II antigen(s), elicited by a non-T, non-B ALL cell line, were characterized by immunofluorescence flow cytofluorometry and ELISA immunofiltration measurements of their immunoreactivity with selected neoplastic hemopoietic cell lines, determination of their complement-dependent cytotoxic activity against isolated peripheral blood B and T lymphocytes and by two-dimensional electrophoretic analysis (isoelectric focusing, SDS-PAGE) of radiolabeled, immunoprecipitated by these antibodies cell surface antigens. Patterns of these immunological reactivities, as well as two-dimensional radioimmunoprecipitation patterns (acidic heavy chain p35 and basic light chain p30) of antigens recognized by these antibodies confirm their anti-MHC class II specificity. One of these antibodies (braFB6; IgG2b) displayed identical pattern of expression on cell lines and cell types as the typical anti-MHC class II antibodies, but immunoprecipitated only a single chain p30 radioiodinated cell surface protein (with two-dimensional pattern close to the beta-chain of MHC class II DR antigen). These properties indicate the ability of braFB6 monoclonal antibody to recognize a nonpolymorphic determinant of DP-MHC class II antigen.
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PMID:Supplementary characteristics of anti-MHC class II monoclonal antibodies elicited by an ALL cell line: immunofluorescence cytofluorometry, C-dependent cytotoxicity, two-dimensional analysis of antigen. 330 87

Nine strains, isolated from leukoplakias or normal mucosa of the oral cavity, and representing the species Candida albicans, C.tropicalis, and Torulopsis glabrata were tested for the capacity to degrade IgA1, IgA2, and S-IgA in liquid cultures. IgA fragments were characterized by SDS-PAGE of culture supernatants in combination with immunoblotting analysis using antibodies specific for heavy chain and light chain determinants. Strains of C.albicans and C.tropicalis were found to express stronger proteolytic activity than a strain of T.glabrata. The three types of IgA were all degraded, alpha-chains being the primary targets. Immunoblotting analysis indicated that divalent fragments corresponding to the deletion of one or both of the Fc alpha constant domains (F(abc)2 alpha or F(ab)2 alpha) were produced. Monovalent half-molecules corresponding to these fragments could also be detected, suggesting that the yeast strains were capable of cleaving inter-alpha-chain disulphide bridges. The possible consequences of yeast-induced degradation for the function of IgA antibodies are discussed.
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PMID:Degradation of IgA1, IgA2, and S-IgA by Candida and Torulopsis species. 332 61

Plasma kallikrein activated spontaneously during the purification of prekallikrein (I) and acetone-activated plasma kallikrein (II) were at pH 7.4 both capable of reducing the capacity of purified human high molecular weight kininogen (HMrK) to function as cofactor in the contact phase activation of factor XII in a crude plasma preparation. At pH 6.8 only I had such an effect. SDS polyacrylamide gel electrophoresis with reduction indicated that both I and II contained kallikrein as a cleaved 'three-chain molecule. I contained in addition a Mr 49,000 fraction reflecting possibly uncleaved heavy chain. The registration of reduced cofactor function of HMrK induced by plasma kallikrein is discussed in view of the assay procedure used.
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PMID:Reduced or unchanged cofactor function of human high molecular weight kininogen induced by human plasma kallikrein. 349 69

Monoclonal and polyclonal antibodies to the variable portions of antigen receptors (anti-idiotypes and anti-idiotopes) are often employed to study the molecular nature and the biological role of these antigen receptors. Such antibodies are operationally defined as those antibodies which bind to a particular immunoglobulin but not to other immunoglobulins of the same class in a radioimmunoassay or ELISA. The monoclonal antibodies 32D1 and 31D1 were initially defined as anti-idiotypic as they recognized an immunoglobulin preparation from the murine B cell lymphoma BCL1, but not other immunoglobulins of the same isotype as assessed by a radioimmunoassay. A potential artifact in defining anti-idiotypic antibodies in this way is the possibility of copurification of antigen and antibody, resulting in the tentative identification of anti-antigen as anti-idiotype. Previous studies have demonstrated that BCL1-IgM is involved in binding of murine leukemia virus (MuLV), and BCL1 immunoglobulin and MuLV-gp70 apparently co-purified as an immune complex. Disruption of the immune complexes with SDS and sucrose gradient purification of the immunoglobulin was adequate to prepare BCL1 immunoglobulin free of gp70 as assessed by radioimmunoassay with the monoclonal anti-gp70 RA3-4A3. This preparation of immunoglobulin was used to show that 31D1 does not bind to BCL1 immunoglobulin, but to the contaminating gp70 in the BCL1 immunoglobulin preparation. However, MAb 32D1 was definitively proven to be anti-idiotypic as it recognized SDS sucrose density gradient purified IgM and immunoisolated heavy chain and light chain from BCL1 immunoglobulin. Several other lymphomas were recognized by mAb 32D1, including the T cell lymphoma UNC1 and the B cell lymphoma Balenlm17. To determine whether mAb 32D1 recognized immunospecific receptors on these lymphoma cell lines immunoprecipitation studies were performed. Immunoisolation and molecular analysis revealed that mAb 32D1 did not recognize the antigen receptor on these two cells, but instead recognized a cell-specific gp70. This observation demonstrates that monoclonal antibodies to known antigens (in this case an anti-idiotype) can crossreact with apparently unrelated molecules. The potential significance of this cross reaction to the antigens recognized by B cell lymphomas is discussed.
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PMID:Molecules recognized by anti-idiotypic monoclonal antibodies to the B cell lymphoma, BCL1. 350 25


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