Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.30.2 (endonuclease)
18,621 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Nine strains of Helicobacter pylori have been isolated exhibiting spontaneous mutations with a loss of catalase activity. Growth characteristics in vitro were unaffected by the mutation showing that catalase is not essential for growth of Helicobacter pylori. Parent strains and mutants could not be distinguished morphologically from each other when compared by electron microscopy. Restriction endonuclease digestion with HindIII, separated in an 0.7% agarose gel in TBE buffer, showed each pair to be highly related to each other. SDS-PAGE separation of proteins from four mutants and parent strains showed that all mutants lacked a 57 kDa protein. The partial N-terminal sequence of this protein shows homology with maize catalase and may be the subunit of the Helicobacter pylori catalase tetramer. It is concluded that catalase negative mutants of Helicobacter pylori occur spontaneously in vitro, but have not yet been observed in vivo. The paucity of such catalase negative strains in clinical specimens could mean that catalase is a virulence factor in vivo that puts mutants at a selective disadvantage.
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PMID:Catalase negative mutants of Helicobacter pylori. 152 35

Familial hypobetalipoproteinemia, a syndrome associated with low plasma cholesterol levels, can be caused by apoB gene mutations. We identified a healthy 42-year-old man whose total plasma cholesterol level was 80 mg/dl. His plasma very low density lipoprotein (VLDL) contained a unique truncated apoB species, apoB-83, in addition to the normal B apolipoproteins, apoB-100 and apoB-48. Virtually no apoB-83 was detectable in his low density lipoprotein (LDL). From the subject's kindred, we identified nine other hypocholesterolemic subjects whose VLDL contained apoB-83. A tendency for cholelithiasis was noted in the apoB-83 heterozygotes, particularly in the older individuals. From the apparent size of apoB-83 on SDS-polyacrylamide gels and its reactivity with apoB-specific monoclonal antibodies, we estimated that it would contain approximately 3700-3800 amino acids. DNA sequencing of apoB genomic clones from two affected individuals revealed that apoB-83 was caused by a C----A transversion in exon 26 of the apoB gene (apoB cDNA nucleotide 11458). This mutation converts Ser-3750 (TCA) into a premature stop codon (TAA) and creates a unique MseI restriction endonuclease site. Thus, a single nucleotide transversion in the apoB gene results in a unique truncated apoB species, apoB-83, and the clinical syndrome of familial hypobetalipoproteinemia.
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PMID:A truncated species of apolipoprotein B, B-83, associated with hypobetalipoproteinemia. 152 80

Citrobacter diversus brain abscess occurred in two infants in Aberdeen, 5 months apart. These are the first reported cases of this condition in the UK since 1976. Restriction endonuclease analysis with SacI enzyme showed blood and CSF isolates from both patients to be identical and different from 10 other clinical isolates of C. diversus and one C. amalonaticus strain. Furthermore, isolates of C. diversus from patients belonged to biotype "d" whereas control isolates were of biotypes "a" or "e". Because both infants attended the same central and peripheral maternity units, this raised suspicions of long term contamination of the hospital environment by this organism. Sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) patterns of whole-cell proteins and immunoblotting with normal human serum were remarkably homogeneous for all 13 C. diversus strains and thus were not useful for typing. However, the only C. amalonaticus strain was clearly differentiated from C. diversus strains by SDS-PAGE. Management of the infants included multiple intravenous antibiotic therapy for 4-6 weeks and repeated computerised tomography (CT) scanning and drainage of the abscess cavity. Both children survived albeit with some minor degree of brain damage.
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PMID:Citrobacter diversus brain abscess: case reports and molecular epidemiology. 156 Apr 49

We have used the polymerase chain reaction to alter transcriptional and translational signals surrounding the hinfIM gene [encoding M.HinfI methyltransferase (MTase)] so as to achieve overexpression in Escherichia coli. The PCR-generated hinfIM gene was subcloned in a high-expression vector under control of the hybrid trp-lac promoter. In addition, the positive retroregulator stem-loop sequence derived from the crystal protein-encoding gene of Bacillus thuringiensis was inserted downstream from hinfIM. Using a similar approach, we have also constructed overproducer clones of a deletion mutant of M.HinfI MTase that has 97 amino acids from the C terminus deleted. The plasmid from the mutant clones is fully protected from HinfI restriction endonuclease digestion. It appears that the functional properties (the recognition and catalytic functions) are encoded within this mutant gene. The overproducer clones yield the wild type (wt) and the mutant enzymes to about 10% of total cellular protein upon induction with 1 mM IPTG. The wt M.HinfI and the mutant MTase were purified to near electrophoretic homogeneity by phosphocellulose, DEAE and gel chromatography. Their monomer sizes by SDS/polyacrylamide-gel electrophoresis are 43 kDa and 31 kDa, respectively, in good agreement with that predicted from the nucleotide sequence. DNA methylation experiments with purified enzymes using single-strand and double-strand M13mp18 DNA substrates indicate that while wt enzyme methylates both forms of DNA substrates, the mutant enzyme appears to preferentially methylate ss DNA substrate.
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PMID:Overproduction, purification and characterization of M.HinfI methyltransferase and its deletion mutant. 156 35

Sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) of whole-cell proteins (WCP), immunoblot analysis and DNA restriction-endonuclease analysis (REA) were applied as potential typing methods to 31 clinically significant strains of Moraxella (Branhamella) catarrhalis, five of which came from a suspected outbreak of nosocomial infection in a respiratory-diseases ward. Twelve of 31 isolates were placed in four groups, each of which contained strains indistinguishable by the three typing techniques used. Each of a further two groups contained two strains, and they were similar by at least one technique; the remaining 15 strains were unique by all three methods. Four of five strains from the suspected outbreak were indistinguishable by SDS-PAGE of WCP, immunoblotting and REA. Results show that SDS-PAGE of WCP, immunoblotting and REA are suitable techniques for characterising M. catarrhalis and that there is a considerable degree of strain heterogeneity. Nosocomial infection with M. catarrhalis may be relatively common and further epidemiological studies with a combination of typing techniques are indicated.
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PMID:Characterisation of hospital isolates of Moraxella (Branhamella) catarrhalis by SDS-PAGE of whole-cell proteins, immunoblotting and restriction-endonuclease analysis. 162 19

Previous papers have reported that the syncytial mutant HSV-1(13)S11 carries three segregable syn mutations and exhibits its altered phenotype in four different cell lines, i.e. HEp-2, VERO, BHK and HEL both at 34 degrees C and 39 degrees C. Those studies have shown that one of three syncytial loci, designated Syn 5, is located in the Bam HI Q fragment spanning map units 0.296-0.317 of the prototype arrangement. Recombinants obtained from marker transfer experiments with donor BamHI Q fragment, have shown that locus Syn 5 is able to induce cell-to-cell fusion in VERO, BHK and HEL but not in HEp-2 cells. In this paper we have characterized the syn mutant HSV-1(13)S11 with regard to plaque morphology, synthesis of viral polypeptides and glycoproteins, thymidine kinase activity and physical map position of locus Syn 5 on the genome. Pertinent to the syn phenotype, earlier papers claimed that two different polypeptides, thymidine kinase (TK) and glycoprotein H (gH), whose genes map in BamHI Q, may be responsible for the fusion activity. Functional studies on the TK of the syn mutant HSV-1(13)S11 indicate that this polypeptide accumulates normally in infected cells and is a fully active enzyme. The other gene product, gH, has been studied with SDS-PAGE and in radioimmunoprecipitation (RIP) experiments using specific monoclonal antibodies. The results indicate that the amount of gH accumulation in the syn mutant-infected cells is greater than its parental strain. However, new marker transfer experiments described here located locus Syn 5 in 663 base pairs between SstI and EcoRI restriction endonuclease sites at the right end of the BamHI Q fragment, where TK gene overlaps in opposite orientation with UL 24 gene. Altogether these results indicate that the Syn 5 locus segregates from the gene specifying gH, to a region encompassing portions of the TK and UL 24 genes, and that the syn mutation does not affect the expression or activity of TK.
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PMID:Phenotypic and genotypic characterization of locus Syn 5 in herpes simplex virus 1. 164 2

Thirty-two isolates of Coxiella burnetii collected from various hosts ranging from arthropods to man were compared by restriction endonuclease (RE) digestion patterns of chromosomal DNA using SDS-PAGE. SDS-PAGE provided better DNA fragment separation than agarose gel electrophoresis and enabled the differentiation of these isolates into six distinct groups on the basis of DNA restriction fingerprints. Two groups of chronic disease isolates could be distinguished, each having unique RE digestion patterns of chromosomal DNA. Three similar but distinct RE digestion patterns were seen among the group of acute disease isolates. Three additional isolates included in this study exhibited a unique RE digestion pattern and also had a unique plasmid type, designated QpDG. DNA-DNA hybridization on selected isolates quantified the relatedness between several groups and supported the classification of these groups as distinct strains.
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PMID:Differentiation of Coxiella burnetii isolates by analysis of restriction-endonuclease-digested DNA separated by SDS-PAGE. 167 52

Large amounts of histones, H1, H2A, H2B, H3, and H4, were observed in total extracts of T4 lymphocytes and derived cell lines infected with the human immunodeficiency virus (HIV) type 1 or type 2. These histones were simply detectable by analysis of crude cellular extracts by polyacrylamide gel electrophoresis in SDS and staining the proteins with Coomassie blue or by immunoblot assays using specific polyclonal antibodies. The histones were found to be localized in the nucleoplasm, bound to low molecular weight (LMW) DNA in the form of nucleosomes. The mechanism responsible for the accumulation of nucleosomes during HIV infection was found to be due to fragmentation of cellular DNA, a mechanism referred to as apoptosis or programmed cell death in which a nuclear endonuclease becomes activated and cleaves DNA at internucleosomal regions. Accordingly, the LMW DNA accumulated in the course of infection was found to have a characteristic pattern of nucleosomal ladder and its accumulation was reduced in the presence of zinc, a known inhibitor of the endonuclease. Routinely in acute HIV infections, the accumulation of nucleosomes was observed at least 24 hr before lysis of infected cells. In a particular HIV-1 infection, in which the first signals of the cytopathic effect (vacuolization of cells and appearance of syncytia) was observed at Days 6-7 whereas maximal virus production occurred at Days 10-17, the accumulation of nucleosomes was at its maximal level already on Day 6 postinfection. In the nucleoplasm of chronically infected cells producing virus but not manifesting a cytopathic effect, no LMW DNA or histones were detectable. These observations indicate that the cytopathic effect of HIV infection is associated with apoptosis. The detection of histones and oligonucleosomal DNA fragments in the nucleoplasm can be used as a convenient marker for chromatin fragmentation during this process.
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PMID:The cytopathic effect of HIV is associated with apoptosis. 168 28

Variation amongst strains of Helicobacter pylori was examined by 35S-methionine-labelled protein SDS-PAGE (Radio-PAGE), immunoblot and DNA fingerprinting techniques. These methods allowed discrimination amongst isolates and showed total correlation. Pig and baboon gastric Helicobacter pylori-like organisms were found to be very similar to Helicobacter pylori by both Radio-PAGE and immunoblotting, whereas a Helicobacter mustelae isolate was markedly different. The HindIII restriction endonuclease digest patterns of Helicobacter pylori DNA illustrated the considerable genomic variation of this organism. The Radio-PAGE and immunoblot typing methods both gave precise identification of Helicobacter pylori strains, but Radio-PAGE was found to give higher resolution and represents a standardised universally applicable fingerprinting method for Helicobacter pylori.
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PMID:Evaluation of fingerprinting methods for identification of Helicobacter pylori strains. 168 29

Ionizing radiation and radiomimetic compounds, such as hydrogen peroxide and bleomycin, generate DNA strand breaks with fragmented deoxyribose 3' termini via the formation of oxygen-derived free radicals. These fragmented sugars require removal by enzymes with 3' phosphodiesterase activity before DNA synthesis can proceed. An enzyme that reactivates bleomycin-damaged DNA to a substrate for Klenow polymerase has been purified from calf thymus. The enzyme, which has a Mr of 38,000 on SDS-PAGE, also reactivates hydrogen peroxide-damaged DNA and has an associated apurinic/apyrimidinic (AP) endonuclease activity. The N-terminal amino acid sequence of the purified protein matches that reported previously for a calf thymus enzyme purified on the basis of AP endonuclease activity. Degenerate oligonucleotide primers based on this sequence were used in the polymerase chain reaction to generate from a bovine cDNA library a fragment specific for the 5' end of the coding sequence. Using this cDNA fragment as a probe, several clones containing 1.35 kb cDNA inserts were isolated and the complete nucleotide sequence of one of these determined. This revealed an 0.95 kb open reading frame which would encode a polypeptide of Mr 35,500 and with a N-terminal sequence matching that determined experimentally. The predicted amino acid sequence shows strong homology with the sequences of two bacterial enzymes that repair oxidative DNA damage, ExoA protein of S. pneumoniae and exonuclease III of E. coli.
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PMID:Isolation of cDNA clones encoding an enzyme from bovine cells that repairs oxidative DNA damage in vitro: homology with bacterial repair enzymes. 170 95


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