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 basis for the markedly altered intracellular binding of [3H]vincristine in a multidrug-resistant variant (DC-3F/VCRd-5L) of Chinese hamster lung cells (DC-3F) was investigated. Binding of [3H]vincristine by protein in cytosol derived from each cell type exhibited a differing requirement for GTP in MgCl2 containing buffer of low-ionic strength. Binding of [3H]vincristine occurred to cytosolic protein derived from both variant and parental DC-3F cells, but after removal of GTP, binding only occurred to cytosolic protein from parental cells regardless of the presence of added GTP. Although binding by cytosolic protein from parental DC-3F cells did not require GTP, the addition of 0.1 mM GTP increased by two-fold the rate and extent of binding. When cytosol from variant and parental DC-3F cells was incubated with low concentrations of [3H]vincristine in high-ionic strength buffer and analyzed by molecular-sieve HPLC, most of the protein binding [3H]vincristine in parentally derived cytosol eluted as Mr 110,000-115,000 daltons, corresponding to that for dimeric tubulin. The same binding species was not detected in cytosol derived from variant cells. However, these same fractions derived with both parental and variant cytosols contained tubulin as shown by SDS-PAGE and immunoblotting. A smaller peak of [3H]vincristine binding and an amount of tubulin equal to that found in later fractions were found in the void volume during the same HPLC elution runs with cytosol from both variant and parental DC-3F cells. Evidence was also obtained for differences between parental and variant DC-3F cells in beta-tubulin isoforms following isoelectric focusing and immunoblotting. Parental-cell cytosol contains a single isoform of beta-tubulin. However, in variant cell cytosol the same isoform and, in addition, three more basic isoforms were found. These alterations in [3H]vincristine binding and in isoform compositions of beta-tubulin in variant versus parental DC-3F cells may have importance in regard to vincristine resistance in DC-3F cells.
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PMID:Altered molecular properties of tubulin in a multidrug-resistant variant of Chinese hamster cells selected for resistance to vinca alkaloids. 304 33

We have isolated a highly enriched preparation of the multienzyme complex which synthesizes deoxyribonucleoside triphosphates (dNTPs) from bacteriophage T4-infected bacteria. By a combination of SDS polyacrylamide gel electrophoresis and assays for specific enzyme activities, we have been able to identify in our final preparation ten different gene products which were previously identified as constituents of this complex, based upon studies with crude preparations. The complex dissociates at high concentrations of NaCl and MgCl2 but is stable under ionic conditions thought to exist in vivo. The purified complex catalyzes the efficient five-step conversion of dCTP to dTTP. Experiments with several T4 mutants have demonstrated that gene products encoded by cd, regA, nrdA, and nrdB are necessary to retain physical integrity of the complex throughout the preparative procedure, while gp44, gp55, and gppseT are not required. We conclude from this evidence that the T4 early gene products which function in dNTP biosynthesis are, in fact, physically linked as a multienzyme complex, and that regA contributes to the integrity of this complex. However, the dNTP-synthesizing complex as we isolate it contains no detectable DNA polymerase, nor have other known replication proteins been detected.
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PMID:T4 phage deoxyribonucleoside triphosphate synthetase: purification of an enzyme complex and identification of gene products required for integrity. 307 39

Neurofilaments were assembled in vitro from the high speed supernatant of mammalian CNS homogenate in a disassembly buffer containing 4-morpholine-ethane sulfonic acid, MgCl2 and EGTA at 4 degrees C in the presence of 4 M glycerol. The assembled neurofilaments were depolymerized by dialysis against the disassembly buffer and repolymerized by the addition of glycerol to the clarified supernatant obtained afer disassembly. The filament assembly reaction was complete in less than 30 s as measured by turbidimetric changes at 415 nm and did not require any added nucleotide. No assembly of filaments was detected when using frozen tissue. The assembled filaments corresponded to the enrichment of neurofilament triplet, the 210,000, 160,000 and 70,000 dalton polypeptides on SDS-polyacrylamide gels and appeared morphologically and immunochemically identical to neurofilaments isolated by axonal flotation methods. These studies demonstrate in vitro assembly of neurofilaments under native conditions which raises the possibility that like microtubules, neurofilaments or a subpopulation of neurofilaments might be in a dynamic state of assembly--disassembly in situ.
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PMID:In vitro assembly and isolation of neurofilaments and microtubules from mammalian CNS. 311 60

The DNA-binding form of the calf uterine androgen receptor (AR) was subjected to limited protease digestion using chymotrypsin, trypsin and a rat prostate cytosol protease. The properties of the generated polypeptide fragments were identified and compared with those of the intact AR. Physicochemical characterization was achieved through sedimentation analysis, gel filtration chromatography and DEAE anion exchange chromatography. Intactness of functional binding domains was evaluated by measuring the retention of steroid- and DNA-binding capacity. Under non-denaturing conditions the intact AR is a highly asymmetrical molecule with a Stokes radius (RS) of 45A, a sedimentation coefficient of 4.3S and a relative molecular mass of 80,000 daltons. This form of AR has an intrinsic binding affinity for DNA and was eluted from DNA-cellulose with 9 mM MgCl2. Chymotrypsin produced a more globular polypeptide (RS: 31A; 3.1S; 41,000 daltons) with a decreased net negative charge. This fragment also displayed DNA-binding affinity but required a higher concentration of MgCl2 (14 mM) for DNA-cellulose elution, indicating an increased affinity for DNA. The observed reduction in molecular size upon chymotrypsin treatment was confirmed when analysed by SDS-polyacrylamide gel electrophoresis after covalently labelling of the AR with [3H]R1881. Rat prostate cytosol contains a protease which is very active in generating an AR polypeptide with an increased affinity for DNA, without changing the AR net negative charge (RS: 33A; 3.7S; 51,000 daltons). The specificity of this protease remained unknown since none of a large number of inhibitors was able to inactivate this enzyme. The fragment generated is different from that obtained with chymotrypsin since significant differences in size as well as in charge were measured. Trypsin treatment generated a much smaller polypeptide (RS: 25A; 2.9S; 30,000 daltons) which had lost its DNA-binding capacity, but not its steroid binding site. This form probably represents the so-called meroreceptor. When intact AR was treated sequentially with prostate cytosol and trypsin, a polypeptide fragment with identical properties was obtained, indicating the spatial separation of two of the proteolytic cleavage sites. These studies provide evidence for the distinct nature of the molecular domains for androgen and DNA interaction on the calf uterine AR.
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PMID:Analysis of steroid- and DNA-binding domains of the calf uterine androgen receptor by limited proteolysis. 330 38

Non-toxic concentrations of various substances were tested for their influence on the gliding motility of Mycoplasma mobile 163K. A significant inhibitory effect on motility was observed with agents acting on nucleic acid synthesis (mitomycin), protein synthesis (puromycin, chloramphenicol), energy metabolism (p-chloromercuribenzoate, iodoacetate) and with compounds reacting with the cytoplasmic membrane or contractile elements (albumin, cholesterol, EDTA, 2-propanol, procain, CaCl2, MgCl2, colchicin and KI). The surface-active compounds Triton X-100, Tego and SDS increased the gliding velocity significantly in some concentrations and incubation periods. The results suggest that the motility of M. mobile depends on a functional cytoplasmic membrane and that cytoskeletal elements are involved in the gliding mechanism.
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PMID:The influence of various substances on the gliding motility of Mycoplasma mobile 163K. 344 49

The proteins from human milk-fat-globule-membrane were radioiodinated, solubilized and analyzed by SDS-polyacrylamide gel electrophoresis. The solubilized milk-fat-globule-membrane preparations contained six major size classes of components with apparent molecular weights of 155, 70, 58, 52, 42, and 39 kilodaltons. The membrane proteins were significantly more accessible to lactoperoxidase-125I in isolated membrane compared with that of whole cream. Major proteins of apparent molecular weights of 155, 70, 58, 52, 42, and 39 kilodaltons were labeled in whole cream and were extracted from the fat-globules membrane with magnesium chloride. Residual cream (after being extracted with MgCl2) showed the loss of the above proteins components. Using an indirect immunoperoxidase staining method and the antibodies to MFGM which immunoprecipitated all the six major glycoprotein components of MFGM, demonstrated their presence on the apical plasma membrane of mammary epithelial cells lining the breast duct in tissue sections. The asymmetric arrangements of proteins in the human milk-fat-globule-membranes, after secretion, is discussed.
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PMID:Investigation into the asymmetric distribution of proteins in human milk-fat-globule-membranes. 351 87

A Ca2+-binding protein named CAB-27 was purified from bovine brain 100,000 g supernatant. The protein has a molecular mass of 27,000 Da as determined by SDS/polyacrylamide-gel electrophoresis and 35,500 Da by sedimentation-coefficient and Stokes-radius analysis. The protein contains about 26% Glx and Asx and 13% basic residues. The acidic nature of the molecule is confirmed by its pI of 4.80. In the presence of 3 mM-MgCl2 and 150 mM-KCl, CAB-27 binds 2.0 mol of Ca2+/mol of protein, with an apparent Kd of 0.2 microM. Ca2+-binding is unaffected by prior incubation of the protein at 80 degrees C for 2 min. Brain contains about 130 mg of CAB-27/kg. Immunoblotting identified CAB-27 in several bovine tissues; it appears to be particularly rich in brain and kidney. In addition, CAB-27 is identified as an inhibitor of bovine pancreas phospholipase A2 in vitro. The inhibitory activity of CAB-27 was 20-fold less potent than lipocortin. On the basis of the Ca2+-binding properties, intracellular concentration and tissue distribution of this protein, we suggest that CAB-27 may be an important intracellular Ca2+ receptor.
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PMID:Purification and characterization of the 27,000 Da calcium-binding protein of bovine brain. 366 33

A Ca2+-binding protein of molecular mass 48 kDa and named 'CAB-48' has been purified from bovine brain 100,000 g supernatant. About 30 mg of CAB-48 was purified from 1 kg of bovine brain. The protein has been characterized with respect to its physical, chemical and Ca2+-binding properties. It has an apparent molecular mass of 48 kDa by SDS/polyacrylamide-gel-electrophoresis and 75.2 kDa from sedimentation-velocity and Stokes-radius data. The acidic nature of the molecule is suggested by its pI of 4.7. In the presence of 3.0 mM-MgCl2 and 150 mM-KCl, CAB-48 binds 1.0 mol of Ca2+/mol of protein with an apparent Kd of 15 microM. A tyrosine protein kinase partially purified from rat spleen catalysed the incorporation of 0.73 mol of phosphate/mol of CAB-48, and phosphoamino acid analysis revealed that phosphorylation of CAB-48 was specific for tyrosine residues.
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PMID:The 48 kDa Ca2+-binding protein of bovine brain. 367 60

This work presents the purification and further characterization of the aldehyde dehydrogenase reconstitutively active in fatty alcohol oxidation, from rabbit intestinal microsomes. Microsomal aldehyde dehydrogenase was solubilized with cholate and purified by using chromatography on 6-amino-n-hexyl-Sepharose and 5'-AMP-Sepharose. The purified enzyme migrated as a single polypeptide band with molecular weight of 60,000 on SDS-polyacrylamide gel. By gel filtration in the presence of detergent, its apparent molecular weight was estimated to be 370,000. In the detergent-free solution, in contrast, it had a much higher molecular weight, indicating its association in forming large aggregates. The pH optimum was 9.0 when pyrophosphate buffer was used. The enzyme was active toward various aliphatic aldehydes with more than three carbons. The Km value for substrate seemed to decrease with increase in the chain length. The microsomal aldehyde dehydrogenase was not affected by disulfiram and MgCl2, which were, in contrast, highly inhibitory towards the activity of the cytosolic aldehyde dehydrogenase separated from intestinal mucosa.
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PMID:Purification of aldehyde dehydrogenase reconstitutively active in fatty alcohol oxidation from rabbit intestinal microsomes. 375 2

Tubulin-tyrosine ligase (TTL), the enzyme responsible for the reversible addition of a tyrosine residue at the carboxyl end of alpha-tubulin, has been purified from porcine brain using a purification scheme based on standard biochemical procedures. The enzyme preparation was nearly homogeneous (purity greater than 95%), was free of tubulin, and could be stored in the presence of glycerol for several months without loss in activity. To develop a more convenient purification of TTL, we have isolated mouse hybridoma cells secreting antibodies to TTL. These monoclonal antibodies recognize TTL not only in brain tissue but also in the liver of various mammals. Monoclonal antibodies isolated from ascites fluid allowed a rapid purification of TTL from a crude brain extract. TTL stayed bound to the immunoaffinity column in 1.5 M NaCl and was eluted with 3 M MgCl2. Highly active TTL was recovered nearly quantitatively at greater than 95% purity and could be stabilized in the presence of glycerol. Glycerol gradient centrifugation, SDS gel electrophoresis and immunoblots identified TTL as a monomeric protein with an apparent polypeptide molecular weight of about 40,000. A one to one complex of TTL with alpha beta-tubulin was observed by gradient centrifugation.
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PMID:Purification of brain tubulin-tyrosine ligase by biochemical and immunological methods. 396 74


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