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)

Six types of plasmid-mediated carbenicillinases can be distinguished on the basis of their substrate profiles, molecular mass isoelectric values and immunological properties. As yet, no structural classification has been attempted for these enzymes at the molecular level. We have isolated the PSE-4 structural gene responsible for carbenicillinase production in Pseudomonas aeruginosa strain Dalgleish and studied its expression in E. coli. A detailed physical map of the cloned fragment and the construction of deletion mutants permitted the precise localization of the PSE-4 structural gene. Various restriction endonuclease fragments known to be flanking or internal to the PSE-4 bla gene were used as DNA probes and tested for homologous sequences in other beta-lactamase genes. A collection of three restriction fragment probes internal or delimiting the PSE-4 structural gene were hybridized with purified plasmid DNA coding for 18 other beta-lactamases. Under high stringency conditions, only the PSE-1, CARB-3 and CARB-4 genes cross-hybridized with PSE-4; while one of the probes tested hybridized solely with CARB-3. Further analysis indicated that the PSE-1, PSE-4, CARB-3 and CARB-4 bla genes are related and could presumably have evolved from a common progenitor.
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PMID:Development of gene probes and evolutionary relationships of the PSE-4 bla gene to plasmid-mediated beta-lactamases of gram-negative bacteria. 278 7

Pseudomonas sp. strain PN-1 is reclassified as Alcaligenes xylosoxidans subsp. denitrificans PN-1. Strain PN-1 is a gram-negative, rod-shaped organism, is motile by means of lateral flagella, is oxidase positive, and does not ferment sugars. Plasmid pCBI, carrying genes for the anaerobic degradation of benzoate in strain PN-1, is 17.4 kilobase pairs in length and is transmissible to a number of denitrifying Pseudomonas aeruginosa and Pseudomonas stutzeri strains. A restriction endonuclease map was constructed.
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PMID:Plasmid pCBI carries genes for anaerobic benzoate catabolism in Alcaligenes xylosoxidans subsp. denitrificans PN-1. 282 51

The egl gene of Pseudomonas solanacearum was cloned on a cosmid and expressed in Escherichia coli. Restriction endonuclease mapping, transposon mutagenesis, and subclone analysis showed that the egl gene was located on a 2.7-kilobase XhoI-SalI P. solanacearum DNA fragment. Immunoabsorption experiments and sodium dodecyl sulfate-polyacrylamide gel electrophoretic analysis showed that the egl gene encodes the 43-kilodalton endoglucanase that is the major excreted endoglucanase of P. solanacearum. In E. coli, the egl gene appeared to be expressed from its own promoter, but its product was restricted to the cytoplasm. The cloned egl gene was mutagenized with Tn5 and used to specifically mutate the chromosomal egl gene of P. solanacearum by site-directed mutagenesis. The resultant mutant was identical to the wild-type strain in production of extracellular polysaccharide and extracellular polygalacturonase as well as several other excreted proteins but produced at least 200-fold less endoglucanase. This mutant strain was significantly less virulent on tomato than the wild-type strain in plant bioassay experiments. Virulence of the endoglucanase-deficient strain was restored to near wild-type levels by complementation in trans with the cloned egl gene, indicating that the egl gene is important but not absolutely required for pathogenesis.
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PMID:Cloning of the egl gene of Pseudomonas solanacearum and analysis of its role in phytopathogenicity. 283 63

The gene encoding the outer membrane phosphate-selective porin protein P from Pseudomonas aeruginosa was cloned into Escherichia coli. The protein product was expressed and transported to the outer membrane of an E. coli phoE mutant and assembled into functional trimers. Expression of a product of the correct molecular weight was confirmed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blot (immunoblot) analysis, using polyclonal antibodies to protein P monomer and trimer forms. Protein P trimers were partially purified from the E. coli clone and shown to form channels with the same conductance as those formed by protein P from P. aeruginosa. The location and orientation of the protein P-encoding (oprP) gene on the cloned DNA was identified by three methods: (i) mapping the insertion point of transposon Tn501 in a previously isolated P. aeruginosa protein P-deficient mutant; (ii) hybridization of restriction fragments from the cloned DNA to an oligonucleotide pool synthesized on the basis of the amino-terminal protein sequence of protein P; and (iii) fusion of a PstI fragment of the cloned DNA to the amino terminus of the beta-galactosidase gene of pUC8, producing a fusion protein that contained protein P-antigenic epitopes. Structural analysis of the cloned DNA and P. aeruginosa chromosomal DNA revealed the presence of two adjacent PstI fragments which cross-hybridized, suggesting a possible gene duplication. The P-related (PR) region hybridized to the oligonucleotide pool described above. When the PstI fragment which contained the PR region was fused to the beta-galactosidase gene of pUC8, a fusion protein was produced which reacted with a protein P-specific antiserum. However, the restriction endonuclease patterns of the PR region and the oprP gene differed significantly beyond the amino-terminal one-third of the two genes.
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PMID:Cloning of the Pseudomonas aeruginosa outer membrane porin protein P gene: evidence for a linked region of DNA homology. 283 40

The hamster elongation factor 2 gene was isolated from genomic libraries of diphtheria toxin- and Pseudomonas aeruginosa exotoxin A-resistant cells containing non-ADP-ribosylatable elongation factor 2, and its structure was determined by a combination of restriction endonuclease mapping and DNA sequence analysis. The entire gene is about 6 kilobases long and has 13 exons. Almost all the introns are about 90-200 bases long, except the first and third, which are about 1 kilobase and 400 bases long, respectively. The first exon is processed just after the initiation codon for translation. The promoter of this gene was also characterized. As this gene contains the mutation conferring resistance to diphtheria toxin and P. aeruginosa exotoxin A, introduction of this gene into mammalian cells results in expression of toxin resistance. Using this characteristic, gene expression by deletion mutants of the 5'-flanking region were examined, and results showed that about 60 base pairs upstream of the TATA sequence were most efficient for expression of the elongation factor 2 gene.
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PMID:Complete nucleotide sequence and characterization of the 5'-flanking region of mammalian elongation factor 2 gene. 283 76

The syrA and syrB genes involved in syringomycin production in Pseudomonas syringae pv. syringae B301D were identified from an EcoRI-pLAFR3 cosmid library and then physically and functionally analyzed in relation to plant pathogenicity. Homologous recombination of the genes required for syringomycin production from cosmids pGX183 (syrA) and pGX56 (syrB), respectively, introduced into nontoxigenic (Tox-) Tn5 mutants W4S2545 and W4S770 resulted in the concomitant restoration of toxin production and full virulence. The disease indices of the Tox+ strains obtained by recombination of the cloned, homologous DNA into the corresponding Tn5 mutant were essentially equivalent to that of strain B301D-R and significantly higher than those of W4S2545 and W4S770. A 12-kilobase (kb) EcoRI fragment from pGX183 was subcloned (i.e., pGX15) and found to contain the sequences necessary for syringomycin production. A map of pGX15 prepared by a combination of restriction endonuclease digestions and Tn5 mutagenesis showed that the syrA sequence was 2.3 to 2.8 kb. Marker exchange of syrA::Tn5 from pGX15 into B301D-R yielded nonpathogenic phenotypes, indicating that syrA is a regulatory gene since it is necessary for both syringomycin production and pathogenicity. The 4.9-kb EcoRI fragment from pGX56 was subcloned (i.e., pGX4) and shown to carry the syrB sequence which was 2.4 to 3.3 kb. Sodium dodecyl sulfate-polyacrylamide gel electrophoretic analysis of protein extracts from B301D-R associated five proteins, ranging from approximately 130,000 to approximately 470,000 in molecular weight, with syringomycin production. The syrA and syrB genes were required for the formation of proteins SR4 (approximately 350,000) and SR5 (approximately 130,000), which are believed to be components of the syringomycin synthetase complex.
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PMID:Physical and functional analyses of the syrA and syrB genes involved in syringomycin production by Pseudomonas syringae pv. syringae. 284 10

14 new transposable phages (TP) were isolated from approx. 200 clinical isolates of Pseudomonas aeruginosa. The frequent occurrence of TP of P. aeruginosa has been confirmed. There are at least two different groups of TP, namely, the group of D3112 and that of B3. The distinctive features of phages belonging to the groups are as follows: 1) low level of DNA-DNA homology (less than 10%), the whole region of homology in phage genomes of different groups being located on right genome end (29-38 kb); only one of phages of the B3 group shows an additional homology with D3112 DNA outside the above mentioned region; 2) a variable DNA is observed on the left end of the B3 group phage genomes and no such DNA is revealed on the left end of genomes of the D3112 group phages; 3) all phages of the B3 group have specific type of interaction with RPL11 plasmid, which distinguish them from phages of the D3112 group; 4) phages belonging to the two groups differ greatly in their growth in cells harbouring pMG7 plasmid which mediates production of PaeR7 endonuclease and in the number of DNA sites sensitive to SalGI, PstI, BglII endonucleases. Since some of the B3 group phage genomes possess BamH1 sites, resistance to this enzyme cannot be regarded as a general characteristics for all TP of P. aeruginosa, as it was earlier proposed. Some aspects of modular hypothesis of bacteriophage evolution concerning, in particular, the ways of module formation are discussed.
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PMID:[Characteristics of phages-transposons of Pseudomonas aeruginosa belonging to 2 groups distinguished by DNA-DNA homology]. 299 Oct 85

PaeI, a new restriction endonuclease from Pseudomonas aeruginosa clinical strain was isolated and characterized. It recognizes and cleaves the sequence 5'-GCATG reduced C-3' generating DNA fragments with 3'-tetranucleotide sticky ends. DNAs of pBR322, SV40 and bacteriophage lambda have one, two and six PaeI recognition sites, respectively. Seventy-two strains of Pseudomonas, Clostridium, Escherichia coli, Shigella, Proteus and Saccharomyces were screened for the presence of site-specific endonucleases. Here we describe the PaeI restriction enzyme found in Pseudomonas aeruginosa; other data will be published elsewhere. Earlier Hinkle and Miller isolated from P. aeruginosa a PaeR7 restriction endonuclease recognizing and cleaving a sequence 5'-C reduced TCGAG-3' (1). Sequence analysis of DNAs cleaved by PaeI shows that the enzyme is the isoschizomer of SphI (2).
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PMID:A site-specific endonuclease from Pseudomonas aeruginosa. 299 50

Plasmid fingerprinting provides a rapid and dependable means of identifying bacterial isolates of the same strain. The stability, wide distribution, and diverse nature and size of extrachromosomal elements make it suitable for virtually all bacterial genera. There are many different procedures available for plasmid screening, and the one chosen depends primarily on the types of organisms to be analyzed. Some procedures are better suited to gram-positive organisms; others are better for visualizing the very large plasmids often seen in Pseudomonas and Rhizobium species. The key to the plasmid fingerprinting technique is agarose gel electrophoresis. In this step of the technique, it is important to differentiate open circular from closed circular forms of plasmids and to recognize the "smile effect." Plasmid fingerprinting can be utilized for epidemiologic studies of both nosocomial and community-acquired infections. The use of restriction endonuclease analysis can greatly enhance the ability of the investigator to differentiate strains that harbor only a single plasmid. Plasmid fingerprinting often provides the only differential characteristic for strains involved in epidemics.
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PMID:Plasmid fingerprinting. A tool for bacterial strain identification and surveillance of nosocomial and community-acquired infections. 299 42

Pseudomonas putida MT53 contains a TOL plasmid, pWW53, that encodes toluene-xylene catabolism. pWW53 is nonconjugative, is about 105 to 110 kilobase pairs (kbp) in size, and differs significantly in its restriction endonuclease digestion pattern and incompatibility group from the archetypal TOL plasmid pWW0. An RP4::pWW53 cointegrate plasmid, pWW53-4, containing about 35 kbp of pWW53 DNA, including the entire catabolic pathway genes, was formed, and a restriction map for KpnI, HindIII, and BamHI was derived. The entire regulated meta pathway genes for the catabolism of m-toluate were cloned into pKT230 from pWW53 on a 17.5-kbp HindIII fragment. The recombinant plasmid supported growth on m-toluate when mobilized into plasmid-free P. putida PaW130. A restriction map of the insert for 10 restriction enzymes was derived, and the locations of xylD, xylL, xylE, xylG, and xylF were determined by subcloning and assaying for their gene products in both Escherichia coli and P. putida hosts. Good induction of the enzymes by m-toluate and m-methylbenzyl alcohol but not by m-xylene was measured in P. putida, but little or no regulation was found in E. coli. The restriction map and the gene order showed strong similarities with published maps of the DNA encoding both the entire meta pathway operon (xylDLEGFJIH) and the regulatory genes xylS and xylR on the archetype TOL plasmid pWW0, suggesting a high degree of conservation in DNA structure for the catabolic operon on the two different plasmids.
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PMID:Evolutionary conservation of genes coding for meta pathway enzymes within TOL plasmids pWW0 and pWW53. 299 36


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