Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:Q07644 (polypeptide)
 72,197 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

When rat liver nuclei are isolated in the presence of the irreversible sulfhydryl-blocking reagent iodoacetamide, digested with DNase I and RNase A, and extracted with 1.6 M NaCl, nuclear envelope (NE) spheres depleted of intranuclear material, as analysed by thin-section electron microscopy, are obtained. Two-dimensional isoelectric focusing (IEF)/SDS-PAGE and non-equilibrium pH gradient electrophoresis (NEPHGE)/SDS-PAGE reveal that the predominant polypeptides are lamins A, B and C. Nuclei isolated in the absence of sulfhydryl blocking reagents yield salt- and nuclease-resistant structures which contain sparse but demonstrable intranuclear material. A number of non-histone polypeptides are seen in addition to the lamins. Nuclei treated with the sulfhydryl cross-linking reagent sodium tetrathionate (NaTT) yield, after exposure to nucleases and 1.6 M NaCl, nuclear matrix-like structures containing an extensive intranuclear network and components of the nucleolus in addition to the NE. Increased amounts of the non-lamin, non-histone polypeptides are recovered with these structures. Subsequent treatment of these NaTT-cross-linked structures with reducing agents in 1.0 M NaCl selectively solubilizes the intranuclear components but leaves the nuclear envelope apparently intact. The lamins remain sedimentable and are virtually absent from the soluble (intranuclear) material. Instead, the major solubilized polypeptides are (a) 68 and 63 kD polypeptides which migrate in the vicinity of lamins B and C, respectively, but are distinguishable from the lamins by immunoblotting and by uni-dimensional peptide mapping; (b) a series of basic 60-70 kD polypeptides (pI greater than 8.0) which are not recognized by anti-lamin antisera; (c) an acidic (pI 5.3) 38 kD polypeptide; and (d) a number of high molecular mass (greater than 100 kD) polypeptides. These observations not only suggest a convenient method for fractionating matrix structures from rat liver nuclei into biochemically and morphologically discrete components, but also identify a subset of major non-lamin, non-histone nuclear polypeptides (comprising approx. 20% of the total nuclear protein) whose intermolecular interactions can be reversibly stabilized apparently by intermolecular disulfide bond formation by NaTT.
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PMID:A subset of non-histone nuclear proteins reversibly stabilized by the sulfhydryl cross-linking reagent tetrathionate. Polypeptides of the internal nuclear matrix. 649 45

The coexistence of densely packed microtubule- and microfilament-based elements in the apex of ciliated epithelial cells, such as the lateral (L) cells of freshwater mussel gill, suggests that with this system it may be possible to define structural connections and interactions that permit integrated cytoskeletal responses to known physiological stimuli. In this study we examine the structure of the L cell apex in detail. The central elements of the elaborate cytoskeleton of the cortex are the basal bodies whose specialized accessory processes are points of integration and the focus of cortical microtubule and microfilament networks. Each basal body supports a cilium and interacts with adjacent basal bodies and with the cell periphery via a dual set of fibre-containing flat trabeculae, both of which are attached to a special organizing centre, the basal foot cap. The distal trabecula is composed of microtubules and the proximal of microfilaments. Connecting the trabeculae, at vertices in the filamentous grids, are core bundles of microfilaments from apical microvilli. In this way, a zig-zag pattern that characterizes microvillar organization at the cell surface is generated. At the cell periphery, the microfilaments from basal foot caps join a peripheral band of microfilaments that underlies the cell border and is associated with four special sites, one in each corner of the cell. Mussel gill epithelial cells contain a polypeptide that resembles actin in its mobility in sodium dodecyl sulphate/10% (w/v) polyacrylamide gels and its affinity for DNase I. Decoration with heavy meromyosin demonstrates that many microfilaments of the L cell apex contain actin, including the microvillar core and peripheral band microfilaments. Actin-associated proteins are also present in these epithelial cells. The actin filaments of the peripheral band are organized to support contraction of the cell border, which would also affect each element of the cortex. This structural complexity, combined with the limited number of modes of interaction between various elements, suggests that the L cell apical cytoskeleton endows the cell with significant positional and morphogenetic information that could be used to compute organellar and cytoskeletal lengths, spacing and changes upon stimulation.
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PMID:The cytoskeleton of the apical border of the lateral cells of freshwater mussel gill: structural integration of microtubule and actin filament-based organelles. 654 55

An isoactin analysis was performed on L-[35S]cysteine labeled BC3H1 cells to determine if these smooth muscle-like cells synthesize vascular smooth muscle actin. Three different NH2-terminal peptides were identified on thin layer electrophoretograms of DNase I-purified and trypsin-digested BC3H1 cell actin. Results obtained from secondary digestion with thermolysin or Staphylococcus aureus V8 protease showed that the most acidic NH2-terminal peptide was derived from vascular smooth muscle alpha-isoactin. Treatment of cell monolayers with serum-free medium caused a 3-fold increase in the level of alpha-isoactin expression and a concomitant decrease in the level of non-muscle beta- and gamma-isoactin. Cell-cell contact was required for induction of alpha-isoactin, and the effects of serum depletion on isoactin expression and cell growth were reversible. The intensity of about 11 out of 500 polypeptide spots on two-dimensional gels of BC3H1 cell polypeptides also was influenced by the culture conditions. The finding that smooth muscle isoactin expression was coupled to cell growth conditions indicate the potential usefulness of BC3H1 cells in studies of isoactin expression and utilization during vascular smooth muscle development.
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PMID:Induction of vascular smooth muscle alpha-isoactin expression in BC3H1 cells. 669 10

Preliminary indications of the occurrence of actin and myosin in crude matrix vesicle preparations have been reported previously. In the present study extracellular matrix vesicles from rat alveolar bone were isolated. They were further purified by a sucrose density gradient. SDS-polyacrylamide gel electrophoresis of the purified vesicles revealed the presence of a polypeptide with a molecular weight of 43 K daltons and with electrophoretic mobility identical to that of blood platelet actin. The limited proteolysis of both 43 K dalton vesicular polypeptide and actin by Staphylococcus aureus-V8-protease revealed three fragments with identical electrophoretic mobility. In addition, the vesicular preparations inhibited the activity of DNase I, a property typical of actin monomers. Filamentous material extracted from matrix vesicles showed ultrastructuraL features of F-actin. Reaction of this material with heavy meromyosin resulted in arrowhead formation, which is characteristic of acto-heavy meromyosin. The occurrence of actin in extracellular matrix vesicles may account for their budding from the osteoblastic plasma membrane, their possible motility in the matrix, and maintenance of the spherical shape.
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PMID:Occurrence of actin-like protein in extracellular matrix vesicles. 681 27

In a previous report [2] we have described a non-histone protein core which could be isolated from Chinese hamster metaphase chromosomes. This core structure maintained the overall morphology of the metaphase chromosome even after removal of all of the histones, together with many of the non-histone proteins and the bulk of the DNA. As part of our work on the characterization of these core structures, we have developed a novel procedure for the isolation of metaphase chromosomes which avoids the use of high pH buffers and hexylene glycol, as well as eliminating the numerous centrifugation and resuspension steps previously employed. Chromosome cores prepared by 2 M NaCl extraction and DNase I digestion from metaphase chromosomes isolated under these more gentle, quasi-physiological conditions, are shown to contain a relatively simple subset of non-histone proteins. One-dimensional SDS-polyacrylamide gel electrophoresis shows two major groups of polypeptides having molecular weights 48 000-52 000 and 65 000-72 000 D respectively, with similarities in mobilities to the nuclear pore complex-lamina polypeptides and tubulins. However, more detailed analysis by two-dimensional gel electrophoresis and peptide mapping has failed to detect these proteins. A 52 000 D polypeptide component of the core is tentatively identified as the intermediate filament protein vimentin. The in vivo significance of chromosome cores is discussed.
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PMID:Chinese hamster metaphase chromosomes isolated under physiological conditions. A partial characterization of associated non-histone proteins and protein cores. 684 Jan 97

A new protein factor that regulates the state of actin polymerization was purified from porcine brain by DNase I-agarose chromatography. The purified protein factor consisted of a 1 : 1 complex of 88,000 dalton polypeptide and actin. When actin was polymerized by salt in the presence of the factor, the steady-state viscosity and the sedimentability were greatly reduced. The extent of the reductions was found to be greater in the presence of Ca2+ than in its absence.
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PMID:A new regulatory protein that affects the state of actin polymerization. 727 60

We describe the use of a gene-targeted random epitope library for the mapping of antigenic determinants. A DNA clone encoding the target antigen was digested randomly with DNase I to generate a population of DNA fragments of different sizes and sequences. After size fractionation, small DNA fragments (100-200 bp) were isolated and cloned into the phage expression vector fUSE2 to form an expression library displaying random polypeptide sequences as fusion proteins at the N terminus of the phage gene III protein. This library, termed a gene-targeted random epitope library to distinguish it from totally random synthetic epitope libraries, was then screened by affinity selection for recombinant phages which were specifically bound by the antibody of interest. Using this approach, we have mapped a monoclonal antibody (mAb)-defined epitope on the bluetongue virus outer capsid protein VP5. This epitope is not accessible on the intact virus surface, but is recognised by the immune system of sheep and cattle during virus infection. Although the example given here utilised a DNA fragment of known sequence and the library was screened for a mAb-defined epitope, the strategy described should be equally applicable to genes of unknown sequence and for screening of epitopes using polyclonal antibodies. The approach can also be extended to identify immunodominant epitope from much more complex genome-targeted random epitope library for virus, bacteria and eukaryotic organisms. Other applications of recombinant phages expressing defined immunodominant epitopes include serodiagnosis and vaccine development.
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PMID:Use of a gene-targeted phage display random epitope library to map an antigenic determinant on the bluetongue virus outer capsid protein VP5. 753 Feb 66

Murine hybridomas were generated to DNA/tight binding proteins complex isolated from the residual nuclear structure following a procedure analogous to that yielding "empty" shells of nuclear envelope. A monoclonal antibody designated 2A8 was selected because of its differential immunostaining of mitotic cells of a synchronized mouse fibroblast cell culture L-929. The target antigen was rendered insoluble by a sequence of extractions of isolated nuclei of diverse cell types with detergents, urea, DNase I and alkali thus reproducing some solubility properties of proteins constituting an operationally defined residual nuclear matrix. The cognate polypeptide was localized on a subset of proteins of M(r) 58-65 kDa, 70 kDa in isolated fibroblast nuclear matrices. The functional implication of the antigen in mitosis-related disassembly-assembly process of the nuclear matrix/envelope was detected. At prophase the antibody decorated the nuclear periphery and nuclear envelope fixed inward filaments. A fibrous network of cytoplasmic localization was stained in metaphase. At anaphase the antigen was dispositioned into peripheral fibrogranular clusters of polar orientation predominantly on one side of the nucleus. Proceeding to telophase a spreading fluorescence was manifested over the entire contour of the nuclear periphery to delineate the reforming nucleus. By immunogold electron microscopy of interphase cells the antigen was identified as evenly distributed in chromatin and interchromatin regions. At initiation of chromosome condensation in mitosis the label was detected predominantly in the chromosomal area.
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PMID:Redistribution of nuclear envelope associated antigen during the mitotic cycle. 761 16

The NS1 polypeptide of minute virus of mice (MVM) is a potent transcriptional activator of the MVM P38 promoter. The minimum region of this promoter required for transactivation has been identified and termed the transactivation region (tar). However, the function of tar and the biochemical steps involved in NS1-mediated transactivation are not well understood. Here we provide evidence that NS1 binds directly and specifically to tar in a strictly ATP-dependent manner. A DNA fragment containing tar was specifically coimmunoprecipitated with purified baculovirus-expressed MVM NS1, using antibodies directed against NS1 amino- or carboxy-terminal peptides. Using this immunoprecipitation assay, we found that the NS1-tar interaction was enhanced approximately 10-fold by ATP, but subsequent incubation at elevated temperatures in the presence, but not the absence, of MgCl2 caused rapid loss of tar binding. This finding suggests that the tar-NS1 complex has a short half-life under assay conditions which favor ATP hydrolysis. Specific binding was efficiently inhibited by self-ligated oligonucleotides containing the core DNA sequence (ACCA)3, but the same nonligated 20- and 21-mer oligonucleotides were unable to compete effectively, indicating that NS1 only binds to its cognate site when this site is presented on DNA fragments of sufficient size. DNase I footprinting experiments performed in the presence of gamma S-ATP revealed that NS1 protects a 43-bp sequence extending asymmetrically from the (ACCA)2 sequence toward the TATA box of the promoter. NS1 footprints obtained at other sites in the MVM genome were similarly large and asymmetric, all extending approximately 31 bp 5' from the core (ACCA)2-3 sequence. Surprisingly, no footprints were obtained in the absence of gamma S-ATP even under low-stringency binding conditions. However, ATP could be omitted from the reactions if NS1 was first incubated with antibodies directed against its 16-amino-acid carboxy-terminal peptide. Since these antibodies probably create intermolecular cross-links, this finding suggests that NS1 may only bind its cognate site efficiently, or perhaps at all, if the transactivator is first induced to form oligomers. From these data, we hypothesize that ATP binding may also induce NS1 to oligomerize and that such assembly is required before the protein can bind effectively to the tar sequence. The functional implications of the NS1-tar interaction will be discussed.
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PMID:Minute virus of mice transcriptional activator protein NS1 binds directly to the transactivation region of the viral P38 promoter in a strictly ATP-dependent manner. 763 87

The transcribing ribonucleoprotein (RNP) complex of human parainfluenza virus type 3 (HPIV-3) requires cellular actin for transcription of viral genome in vitro (De, B. P., Lesoon, A., and Banerjee, A. K. (1991) J. Virol. 65, 3268-3275). In this communication, we have studied the interactions between different molecular forms of actin and the RNP of HPIV-3 to understand the role of actin in mRNA synthesis. We demonstrate that both polymeric and monomeric forms of actin (obtained by DNase I treatment) bind strongly to the RNP at 100 mM KCl concentration (polymerizing buffer). The binding was virtually abolished at zero KCl concentration (depolymerizing buffer). Isolation of the RNP-actin complex and subsequent use in a transcription reaction showed that the bound actin alone was sufficient for mRNA synthesis in vitro. Interestingly, the DNase I-arrested monomeric form of actin failed to activate mRNA synthesis, indicating a requirement of polymerization of the bound actin during HPIV-3 transcription. Electron microscopic analyses revealed that a drastic structural modification of the RNP occurred because of the polymerization of actin from a loosely coiled and irregular structure to a condensed and flexible structure. Activation of transcription was observed also with poly-L-glutamic acid, a highly acidic polypeptide. However, unlike cellular actin, poly-L glutamic acid was able to activate only 10% of the input RNP. These results suggest that cellular actin activates HPIV-3 transcription by polymerizing specifically on the RNP complex. This event results in an alteration of the RNP structure that enhances its suitability for efficient transcription. The acidic domain of actin may play an important role in this process.
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PMID:Role of cellular actin in human parainfluenza virus type 3 genome transcription. 768 Jun 47


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