Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
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With the use of two-dimensional gel electrophoresis, the proteins present in a transformation-proficient B. subtilis strain were compared with those present in an isogenic, recombination-deficient strain carrying the recE4 mutation. One protein (molecular weight 45 kD, iso-electric point 5.4) was found to be virtually absent in the recE4 strain. This 45 kD protein is a prominent protein predominantly present in the competent fraction of a competent culture. The synthesis of the protein is substantially stimulated by irradiation with ultraviolet light or treatment with mitomycin C and, to a lesser extent, by treatment with nalidixic acid. Since the protein is also observed in a strain cured for SP beta and carrying non-inducible PBS X, it is unlikely that this protein is a gene product specified by one of these prophages usually present in B. subtilis strain 168. Based on these results we conclude that the 45 kD protein is involved in recombination in B. subtilis.
Mol Gen Genet 1982
PMID:Transformation of Bacillus subtilis competent cells: identification of a protein involved in recombination. 681 34

Surface membrane immunoglobulin from MOPC-315 plasmacytoma cells (smM315) was isolated by nonionic detergent lysis of radioiodinated cells and affinity chromatography on Dnp-aminohexyl-Sepharose 4B. Verification of the solubilized molecule as an integral membrane protein, distinct from secreted MOPC-315 IgA (M315) was accomplished by NaDodSO4-PAGE, charge-shift electrophoresis and molecular sieve gel filtration with NP-40 and deoxycholate. smM315 was compared to reduced and alkylated monomeric secreted immunoglobulins from MOPC-315, MOPC-460, and XRPC-25 by quantitative affinity chromatography (QAC) using two differently substituted Dnp-aminohexyl-Sepharose 4B resins. Unique patterns of cross-reactivity of all secreted myeloma proteins were independently established with a competitive hapten inhibition assay using 125I-Dnp26BSA as the precipitating probe. After derivation with dinitrobenzylsulfonate, Dnp-aminohexyl-Sepharose 4B was modified with succinic anhydride which, with the inclusion of 0.03% Doc in a PBS and 0.1% NP-40 buffer, prevented nonhapten specific protein-matrix interactions during QAC. Dissociation constants determined by QAC for three ligands, (dinitrophenyl-glycine, trinitrophenyl-amino-caproate and tetramethylrhodamine) were essentially the same for smM315 and M315. Both of the other nitrophenyl binding IgA myelomas had distinct and significant differences in dissociation constants. Thus, for a differentiated antibody secreting cell which has undergone a heavy chain class switch, such as MOPC-315, the cell surface immunoglobulin has an identical ligand binding active-site as the secreted immunoglobulin.
Mol Immunol 1983 Jan
PMID:Determination of dissociation constants and ligand specificity of detergent solubilized surface membrane immunoglobulin A from MOPC-315. 685 76

Ninety-five percent of the neurons in the corpus striatum of the rat are medium spiny projection neurons, which contain tachykinins such as substance P, neurokinin A, and neurokinin B and the opiate peptides, enkephalin and dynorphin. The remaining 5% consist of interneurons, of which a small but significant proportion are cholinergic. The influence of these cholinergic interneurons on the neuropeptidergic projection systems in the striatum is poorly understood at this time. The present study explores the relationship between cholinergic receptor activation or muscarinic blockade on striatal neuropeptide gene expression. Adult male Sprague-Dawley rats were treated chronically either with a cholinergic agonist (physostigmine: 0.5 mg/kg/3 x day), a muscarinic antagonist (scopolamine HCl: 0.4 mg/kg/3 x day), or vehicle (PBS: 0.1 ml/100 g) administered for 6 days (s.c.). In situ hybridization was performed with probes directed against mRNAs for beta-preprotachykinin (a transcript containing substance P, neurokinin A, and other tachykinins), neurokinin B and preproenkephalin. Physostigmine administration resulted in a 12% decrease in the dorsolateral caudate-putamen and a 27% increase in the core of the nucleus accumbens in substance P/neurokinin A mRNA; and a 29% increase in the caudate-putamen and an 11% increase in the core of the nucleus accumbens in preproenkephalin mRNA levels. Scopolamine treatment resulted in a 28% and 48% decrease, respectively, in the caudate-putamen and in the shell of the nucleus accumbens in substance P/neurokinin A mRNA levels. Neurokinin B mRNA levels were increased by 50% in the shell of the accumbens after scopolamine. Preproenkephalin mRNA levels increased by 24% in the caudate-putamen and decreased by 20% in the core of the nucleus accumbens. From these results we tentatively conclude that cholinoceptive neuropeptidergic neurons are segregated along dorsoventral and mediolateral axes in the striatum, thus giving rise to non-homogenous responses upon cholinergic receptor activation or muscarinic blockade.
Brain Res Mol Brain Res 1995 Jun
PMID:Cholinergic regulation of tachykinin- and enkephalin-gene expression in the rat striatum. 763 70

The coating of the wells of microtiter plates with cobrotoxin in Tris (pH 9.8) or PBS (pH 7.2) buffer was assessed by enzyme-linked immunoassay (ELISA). It was found that the poor binding in neutral buffer was improved by adding glutaraldehyde (GA), and the bound amount reached the same extent as that measured with alkali buffer. The optimal concentration of GA was approx. 0.02%. A decrease in optical density was observed with GA concentration higher than 0.02%. This may result from the modification of cobrotoxin by GA, which induced a decrease in the antigenicity of the cobrotoxin as revealed by competitive immunoassay. This study implies that the coating of physiological samples with GA for ELISA may be carried out with physiological buffers without the need to change the buffer to one of alkaline pH.
Biochem Mol Biol Int 1994 Sep
PMID:Improvement in the binding of cobrotoxin to microtiter plates by glutaraldehyde at neutral pH. 784 51

The morphology of the inner cell mass (ICM) cells and the proportion of dead ICM cells in frozen-thawed bovine preimplantation embryos were investigated by differential fluorochrome staining. Embryos at the blastocyst stage of development were frozen and thawed by two different techniques (three-step and one-step) in two different basic salt solutions (PBS and TCM 199) containing 1.36M glycerol. After thawing and glycerol removal, embryos were co-cultured in a cumulus cells monolayer in TCM 199 for 48 hr (morula) or 24 hr (blastocysts). Differential cell counts of the ICM and trophectoderm were then done using differential fluorochrome staining. Overall, there was no significant difference in the viability of embryos frozen in the two basic salt solutions. Low proportions of dead ICM cells were observed in embryos frozen at the morula stage in both PBS (19.1%) or TCM 199 (18.0%). However, blastocyst stage embryos frozen by the three-step technique had a higher (P < 0.05) proportion of dead ICM cells in TCM 199 (37.7%) than in PBS (18.2%). Blastocysts frozen by the one-step technique had a higher (P < 0.05) proportion of dead ICM cells (42.2%) than those frozen by the three-step technique (18.2%), regardless of basic salt solutions. Results indicate that freezing and thawing damages ICM cells in morphologically normal embryos and that the degree of damage depended on the basic salt solution and the freezing method.
Mol Reprod Dev 1994 Mar
PMID:Effects of freezing of bovine preimplantation embryos derived from oocytes fertilized in vitro on survival of their inner cell mass cells. 818 31

The 5' region of HIV-1 RNA contains functional elements involved in key steps of the retroviral cycle, such as genomic RNA transcription, splicing, translation, dimerization or initiation of reverse transcription. In the present work, we investigated the conformation of the first 500 nucleotides covering the RNA leader and the 5' gag coding sequences of HIV-MAL, using chemical probing. We provide detailed information on almost each nucleotide at one of their Watson-Crick positions and on position N-7 of purines. Experiments were conducted on two in vitro transcribed RNA fragments (1 to 707 and 1 to 311). A secondary structure model was derived by combining the experimental data, computer predictions and sequence comparison. Under conditions favoring dimerization (high salt concentration), HIV-1 RNA folds into independent structural domains that can be related to defined functional regions. The first domain corresponds to TAR forming a stable stem-loop. Intrinsic structural features are found to stabilize the TAR hairpin loop. The second domain (nucleotides 56 to 299) contains the PBS sequence, which is located in a stable subdomain constrained by a four stem junction (nucleotides 139 to 218). Although the MAL isolate has an insertion near the PBS, probably resulting from the duplication of a 23-nucleotide sequence, the structural organization of this subdomain is conserved in all other HIV-1 isolates. The third domain (nucleotides 300 to 404) contains the splice donor site, packaging and dimerization elements and the AUG initiation codon of gag. A major result is the structural versatility of this region. Two mutually exclusive structures, both equally in agreement with probing data, could modulate the different functions involving this domain. The reduced accessibility of the gag translational initiation site possibly accounts for the low efficiency of the in vitro translation of the dimer. Finally, the 5' gag coding sequences form a metastable domain.
J Mol Biol 1993 Jan 20
PMID:Functional sites in the 5' region of human immunodeficiency virus type 1 RNA form defined structural domains. 842 53

The time of activation of the embryonic genome (maternal-embryonic transition) in equine embryos was investigated by assessing incorporation of 3H-uridine and nucleolar development. In Experiment 1, embryos were recovered from the oviduct (n = 15) and the uterus (n = 3). Recovered embryos were assessed for morphologic development and quality score. Recovered embryos with less than 8 cells (two cells, n = 4; four cells, n = 5; five cells, n = 2) were incubated with 3H-uridine (560 microCi/ml) for 10 hr, while eight-cell embryos (n = 2), morulae (n = 2), and blastocysts (n = 3) were incubated with 280 microCi/ml for 0.5-1 hr. At the end of incubation, embryos were washed twice in PBS with 10% FBS and incubated for 30 min with 2.5 mg/ml of unlabelled uridine. Embryos were spread onto glass slides, dipped into emulsion, and exposed for 8 d, then developed and counterstained with Giemsa and propidium iodide. Embryos at the blastocyst, morula, eight-cell, and five-cell stages incorporated 3H-uridine into their cell nuclei as detected by autoradiography. In a second experiment, nucleologenesis in equine embryos was examined by transmission electron microscopy. Nucleoli or nucleolar precursors were found in 12 of 23 embryos examined. Most embryos in the four- to six-cell stage (n = 7) had nucleolar precursor bodies (npb) consisting of homogeneous fibrillar structures. Two five- to six-cell embryos also possessed reticulated nucleoli with both fibrillar and granular components as did all eight-cell embryos (n = 3). Nucleoli in one morula and one blastocyst were reticulated with prominent granular components, fibrillar components, and apparent fibrillar centers. These results indicate that incorporation of 3H-uridine and the formation of functional nucleoli with typical fibrillar and granular components occurs between the four- to eight-cell stage in equine embryos.
Mol Reprod Dev 1995 Nov
PMID:Initiation of transcription and nucleologenesis in equine embryos. 857 43

Cu/Zn-superoxide dismutase (Cu/Zn-SOD) has been shown to modulate the autoxidation of a variety of phenoic compounds, including 1,4-hydroquinone (HQ), a benzene-derived metabolite. The acceleration of autoxidation of HQ by Cu/Zn-SOD results in the production of 1,4-benzoquinone (BQ). It has been proposed that the chemical mechanism involved in the Cu/Zn-SOD-catalyzed autoxidation of HQ may be occur through either its conventional activity as a superoxide:superoxide oxidoreductase or as a semiquinone:superoxide oxidoreductase. However, Cu/Zn-SOD-accelerated oxidation of HQ has not been resolved experimentally. In this study, with ESR spectroscopy we investigated further the chemical reactions involved in the SOD-accelerated oxidation of HQ. In phosphate-buffered saline (PSB), HQ underwent a slow autoxidation to BQ, which was accelerated by Cu/Zn-SOD, Mn-SOD, or Fe-SOD with similar efficiency. In contrast, among free metals, only Cu(II) strongly mediated the oxidation of HQ to BQ. Mn(II) exhibited a slight capacity to oxidize HQ, whereas neither FE(II) nor FE(III) was capable of modulating the autoxidation of HG. The presence of either form of SOD also dramatically enhanced the formation of semiquinone anion radicals SQ-. from HQ. The SOD-accelerated oxidation of HQ was also accompanied by the generation of H202. In PBS containing bovine serum albumin (BSA) (PBS/BSA), HQ did not undergo autoxidation to SQ-., and as such the presence of SOD was unable to induce the formation of either SQ-. or BQ or the consumption of O2. The addition of 10 microM BQ to HQ (100 or 1000 microM) in PBS/BSA resulted in the formation of SQ-. and initiated a slow rate of oxidation of HQ to BQ. In this case, the presence of Cu/Zn-SOD strongly accelerated the oxidation of HQ to SQ-. and BQ and the utilization of O2. Furthermore, the enhancement by Cu/Zn-SOD of the generation of SQ-. or BQ from HQ in PBS/BSA was extensively inhibited under anaerobic conditions. The enhancement of SQ-. generation from HQ by all three forms of SOD does not support the possibility that Cu/Zn-SOD can oxidize SQ-. to BQ. Taken together, this study demonstrates that unlike free copper, Cu/Zn-SOD does not directly interact with HQ to cause its oxidation to BQ. Rather, the autoxidation of HQ to SQ-. is a prerequisite for the enhancing capacity of Cu/Zn-SOD, and the dismutation of superoxide anion radicals generated from the SQ-. in the presence of O2 appears to be the underlying mechanism responsible for the enhancement by Cu/Zn-SOD of the oxidation of HQ.
Mol Pharmacol 1996 Mar
PMID:Role of Cu/Zn-superoxide dismutase in xenobiotic activation. I. Chemical reactions involved in the Cu/Zn-superoxide dismutase-accelerated oxidation of the benzene metabolite 1,4-hydroquinone. 864 79

The subunit structure of soluble goat hepatic lectin was studied by determining molecular weight under nondenaturing conditions by gel filtration and denaturing conditions by SDS PAGE. Affinity purified lectin was subjected to HPLC on asahipack column equilibrated with 10 mM Tris-HCl pH 7.5, containing 1 mM CaCl2, 1mM 2-mercaptoethanol and 0.1M NaCl. The lectin was eluted under single peak at retention time of 12 min. corresponding to molecular weight of 38Kd. On SDS-PAGE in the presence and absence of 2-mercaptoethanol protein moved as single band with Rm 0.45, which corresponds to molecular weight of 20 Kd. The results suggest that soluble goat hepatic lectin is a dimmer of identical subunits which are linked together by noncovalent interactions. The interaction of monoclonal antibodies raised against soluble goat hepatic lectin (MGHL/20) with hemagglutinin from different species as sheep, human, rat, bovine and chicken was studied in PBS by solid phase binding assay. MGHL/20 showed 29.89% binding with these lectin. However no binding was found with Ca++ dependent membrane bound lectin. These results indicate that soluble goat hepatic lectin possesses antigenic structural relationship with soluble 14 K lectin family.
Biochem Mol Biol Int 1996 Aug
PMID:Subunit structure of Ca++ dependent soluble goat hepatic lectin: evidence that it has antigenic structural relationship with soluble 14K lectin family. 886 13

Alveolar epithelial injury occurs universally in common respiratory illnesses associated with diffuse lung damage. After alveolar injury, type II cells proliferate and reestablish epithelial integrity, thereby restoring normal lung structure and function. However, the regulation of type II cell proliferation and alveolar epithelial repair is poorly understood. Hepatocyte growth factor/scatter factor (HGF/SF) is a heparin-binding growth factor that has been shown to be mitogenic for cultured alveolar type II cells. In this study, we determined the effect of intratracheal instillation of rhHGF/SF on type II cell proliferation in vivo. To quantify the alveolar type II cell proliferative response, we developed a double-label immunohistochemical technique to detect replicating alveolar type II cells in formalin-fixed lung sections that utilized the identification of proliferating cells by bromodeoxyuridine (BrdUrd) incorporation into DNA and alveolar type II cells by 3F9 immunoreactivity. BrdUrd detection was optimized by enzymatic antigen recovery and silver intensification of the horseradish peroxidase reaction product. Intratracheal instillation of rhHGF/SF induced a time- and dose-dependent increase in type II cell proliferation. The type II cell labeling index increased to 12.3 +/- 6.0% 48 h after 1.0 mg/kg rhHGF/SF administration, compared with 2.6 +/- 0.9% after PBS instillation. To compare the normal type II cell reparative response with the level of proliferation after exogenous rhHGF/SF administration, we measured the specific alveolar type II cell labeling index in rat lung sections obtained from animals exposed to hyperoxia for 50 h and then allowed to recover in room air. After 1 day of recovery, the alveolar type II cell labeling index was 0.45 +/- 0.2%. The specific labeling index increased to 5.4 +/- 1.3% at 2 days and then declined to 0.31 +/- 0.16% 5 days after hyperoxia exposure. In animals not exposed to hyperoxia, the alveolar type II cell labeling index was 0.6 +/- 0.14%. These studies demonstrated that intratracheal instillation of rhHGF/SF promoted alveolar type II cell proliferation in vivo. The maximal level of type II cell proliferation after rhHGF/SF administration was more than twice that reached during recovery from hyperoxia exposure. Thus, intratracheal instillation of HGF/SF may provide a potential strategy to promote type II cell proliferation and augment alveolar epithelial repair after lung injury.
Am J Respir Cell Mol Biol 1996 Nov
PMID:Intratracheal administration of hepatocyte growth factor/scatter factor stimulates rat alveolar type II cell proliferation in vivo. 891 64


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