A mechanism of carbapenem resistance due to a new insertion element (ISPa133) in Pseudomonas aeruginosa

This study explored the evolutionary mechanism by which the clinical isolate PA110514 yields the imipenemresistant derivative PA116136. Both isolates were examined by PFGE and SDS-PAGE, which led to the identification of a new insertion sequence, ISPa133. This element was shown to have distinct chromosomal locations in each of the original isolates that appeared to explain the differences in imipenem susceptibilty. In strain PA110514, ISPa133 is located 56 nucleotides upstream of the translational start codon, which has no effect on expression of the porin OprD. However, in strain PA116136 ISPa133 it is located in front of nucleotide 696 and, by interrupting the coding region, causes a loss of OprD expression, thus conferring imipenem resistance. In vitro experiments mimicking the natural conditions of selective pressure yielded imipenem-resistant strains in which ISPa133 similarly interrupted oprD. A mechanism is proposed whereby ISPa133 acts as a mobile switch, with its position in oprD depending on the degree of selective pressure exerted by imipenem

A mechanism of carbapenem resistance due to a new insertion element (ISPa133) in Pseudomonas aeruginosa

This study explored the evolutionary mechanism by which the clinical isolate PA110514 yields the imipenemresistant derivative PA116136. Both isolates were examined by PFGE and SDS-PAGE, which led to the identification of a new insertion sequence, ISPa133. This element was shown to have distinct chromosomal locations in each of the original isolates that appeared to explain the differences in imipenem susceptibilty. In strain PA110514, ISPa133 is located 56 nucleotides upstream of the translational start codon, which has no effect on expression of the porin OprD. However, in strain PA116136 ISPa133 it is located in front of nucleotide 696 and, by interrupting the coding region, causes a loss of OprD expression, thus conferring imipenem resistance. In vitro experiments mimicking the natural conditions of selective pressure yielded imipenem-resistant strains in which ISPa133 similarly interrupted oprD. A mechanism is proposed whereby ISPa133 acts as a mobile switch, with its position in oprD depending on the degree of selective pressure exerted by imipenem

A mechanism of carbapenem resistance due to a new insertion element (ISPa133) in Pseudomonas aeruginosa

This study explored the evolutionary mechanism by which the clinical isolate PA110514 yields the imipenemresistant derivative PA116136. Both isolates were examined by PFGE and SDS-PAGE, which led to the identification of a new insertion sequence, ISPa133. This element was shown to have distinct chromosomal locations in each of the original isolates that appeared to explain the differences in imipenem susceptibilty. In strain PA110514, ISPa133 is located 56 nucleotides upstream of the translational start codon, which has no effect on expression of the porin OprD. However, in strain PA116136 ISPa133 it is located in front of nucleotide 696 and, by interrupting the coding region, causes a loss of OprD expression, thus conferring imipenem resistance. In vitro experiments mimicking the natural conditions of selective pressure yielded imipenem-resistant strains in which ISPa133 similarly interrupted oprD. A mechanism is proposed whereby ISPa133 acts as a mobile switch, with its position in oprD depending on the degree of selective pressure exerted by imipenem

A mechanism of carbapenem resistance due to a new insertion element (ISPa133) in Pseudomonas aeruginosa

This study explored the evolutionary mechanism by which the clinical isolate PA110514 yields the imipenemresistant derivative PA116136. Both isolates were examined by PFGE and SDS-PAGE, which led to the identification of a new insertion sequence, ISPa133. This element was shown to have distinct chromosomal locations in each of the original isolates that appeared to explain the differences in imipenem susceptibilty. In strain PA110514, ISPa133 is located 56 nucleotides upstream of the translational start codon, which has no effect on expression of the porin OprD. However, in strain PA116136 ISPa133 it is located in front of nucleotide 696 and, by interrupting the coding region, causes a loss of OprD expression, thus conferring imipenem resistance. In vitro experiments mimicking the natural conditions of selective pressure yielded imipenem-resistant strains in which ISPa133 similarly interrupted oprD. A mechanism is proposed whereby ISPa133 acts as a mobile switch, with its position in oprD depending on the degree of selective pressure exerted by imipenem

A mechanism of carbapenem resistance due to a new insertion element (ISPa133) in Pseudomonas aeruginosa

This study explored the evolutionary mechanism by which the clinical isolate PA110514 yields the imipenemresistant derivative PA116136. Both isolates were examined by PFGE and SDS-PAGE, which led to the identification of a new insertion sequence, ISPa133. This element was shown to have distinct chromosomal locations in each of the original isolates that appeared to explain the differences in imipenem susceptibilty. In strain PA110514, ISPa133 is located 56 nucleotides upstream of the translational start codon, which has no effect on expression of the porin OprD. However, in strain PA116136 ISPa133 it is located in front of nucleotide 696 and, by interrupting the coding region, causes a loss of OprD expression, thus conferring imipenem resistance. In vitro experiments mimicking the natural conditions of selective pressure yielded imipenem-resistant strains in which ISPa133 similarly interrupted oprD. A mechanism is proposed whereby ISPa133 acts as a mobile switch, with its position in oprD depending on the degree of selective pressure exerted by imipenem

A mechanism of carbapenem resistance due to a new insertion element (ISPa133) in Pseudomonas aeruginosa

This study explored the evolutionary mechanism by which the clinical isolate PA110514 yields the imipenemresistant derivative PA116136. Both isolates were examined by PFGE and SDS-PAGE, which led to the identification of a new insertion sequence, ISPa133. This element was shown to have distinct chromosomal locations in each of the original isolates that appeared to explain the differences in imipenem susceptibilty. In strain PA110514, ISPa133 is located 56 nucleotides upstream of the translational start codon, which has no effect on expression of the porin OprD. However, in strain PA116136 ISPa133 it is located in front of nucleotide 696 and, by interrupting the coding region, causes a loss of OprD expression, thus conferring imipenem resistance. In vitro experiments mimicking the natural conditions of selective pressure yielded imipenem-resistant strains in which ISPa133 similarly interrupted oprD. A mechanism is proposed whereby ISPa133 acts as a mobile switch, with its position in oprD depending on the degree of selective pressure exerted by imipenem

A mechanism of carbapenem resistance due to a new insertion element (ISPa133) in Pseudomonas aeruginosa

This study explored the evolutionary mechanism by which the clinical isolate PA110514 yields the imipenemresistant derivative PA116136. Both isolates were examined by PFGE and SDS-PAGE, which led to the identification of a new insertion sequence, ISPa133. This element was shown to have distinct chromosomal locations in each of the original isolates that appeared to explain the differences in imipenem susceptibilty. In strain PA110514, ISPa133 is located 56 nucleotides upstream of the translational start codon, which has no effect on expression of the porin OprD. However, in strain PA116136 ISPa133 it is located in front of nucleotide 696 and, by interrupting the coding region, causes a loss of OprD expression, thus conferring imipenem resistance. In vitro experiments mimicking the natural conditions of selective pressure yielded imipenem-resistant strains in which ISPa133 similarly interrupted oprD. A mechanism is proposed whereby ISPa133 acts as a mobile switch, with its position in oprD depending on the degree of selective pressure exerted by imipenem

Insertion of a malE B-Galactosidase fusion protein into the envelope of Escherichia coli disrupts biogenesis of outer membrane proteins and processing of inner membrane proteins

The synthesis of a membrane-bound MalE ,B-galactosidase hybrid protein, when induced by growth of Escherichia coli on maltose, leads to inhibition of cell division and eventually a reduced rate of mass increase. In addition, the relative rate of synthesis of outer membrane proteins, but not that of inner membrane proteins, was reduced by about 50%o. Kinetic experiments demonstrated that this reduction coincided with the period of maximum synthesis of the hybrid protein (and another maltose-inducible protein, LamB). The accumulation of this abnormal protein in the envelope therefore appeared specifically to inhibit the synthesis, the assembly of outer membrane proteins, or both, indicating that the hybrid protein blocks some export site or causes the sequestration of some limiting factor(s) involved in the export process. Since the MalE protein is normally located in the periplasm, the results also suggest that the synthesis of periplasmic and outer membrane proteins may involve some steps in common. The reduced rate of synthesis of outer membrane proteins was also accompanied by the accumulation in the envelope of at least one outer membrane protein and at least two inner membrane proteins as higher-molecular-weight forms, indicating that processing (removal of the N-terminal signal sequence) was also disrupted by the presence of the hybrid protein. These results may indicate that the assembly of these membrane proteins is blocked at a relatively late step rather than at the level of primary recognition of some site by the signal sequence. In addition, the results suggest that some step common to the biogenesis of quite different kinds of envelope protein is blocked by the presence of the hybrid protein.

A chromosomal insertion toolbox for promoter probing, mutant complementation and pathogenicity studies in Ralstonia solanacearum

We describe here the construction of a delivery system for stable and directed insertion of gene constructs in a permissive chromosomal site of the bacterial wilt pathogen Ralstonia solanacearum. The system consists of a collection of suicide vectors the Ralstonia chromosome (pRC) series that carry an integration element flanked by transcription terminators and two sequences of homology to the chromosome of strain GMI1000, where the integration element is inserted through a double recombination event. Unique restriction enzyme sites and a GATEWAY cassette enable cloning of any promoter::gene combination in the integration element. Variants endowed with different selectable antibiotic resistance genes and promoter::gene combinations are described. We show that the system can be readily used in GMI1000 and adapted to other R. solanacearum strains using an accessory plasmid. We prove that the pRC system can be employed to complement a deletion mutation with a single copy of the native gene, and to measure transcription of selected promoters in monocopy both in vitro and in planta. Finally, the system has been used to purify and study secretion type III effectors. These novel genetic tools will be particularly useful for the construction of recombinant bacteria that maintain inserted genes or reporter fusions in competitive situations (i.e., during plant infection).

Reação de bis-inserção de 1,2-difenilacetileno na ligação Pd-C de ciclometalados

The present paper deals with the bis-insertion reactions of 1,2-diphenylacetylene into Pd-C bond of the cyclopalladated complexes [Pd(dmba)(µ-NCO)]2 (1) and [Pd(dmba)(MeCN)2](NO3) (2) (dmba = N,N-dimethylbenzylamine, MeCN = acetonitrile). Two new complexes [Pd{PhC=CPh-CPh=CPhC6H4CH2N(CH 3)2}(NCO)] (3) and [Pd{PhC=CPh-CPh=CPhC6H4CH2N(CH 3)2}(NO3 )] (4) were obtained and characterized by IR and NMR spectroscopy and elemental analysis.

Les N-tosyloxycarbamates : une nouvelle source de nitrènes métalliques pour la réaction d'insertion de liens carbone-hydrogène

Thèse numérisée par la Division de la gestion de documents et des archives de l'Université de Montréal

A carbene insertion approach to functionalised poly(ethylene oxide)-based gels

Carbenes photogenerated from the novel bisdiazirine, 1, 3-bis(3-(trifluoromethyl)diazirin-3-yl) benzene 1, have been applied successfully to cross-linking of mono-methyl poly(ethylene oxide) (MePEO5000) in the presence of dichloromethane, leading to the simultaneous incorporation of alkylhalide functionalities. The PEO-based gels swell in a wide range of solvents with polarity index values varying from 3.1 to 9.0. Reaction of the alkylhalide functionalities present in the gels with 4-phenylazophenol provided loading capacities of up to 0.20 mmol g(-1) and demonstrated the potential of these materials for gel-phase synthesis applications. (C) 2008 Elsevier Ltd. All rights reserved.

Synthesis of hyperbranched poly(aryl ether)s via carbene insertion processes

Homopolymerization of alkylarylcarbenes derived from diazirine monomers that featured benzyl alcohol or phenol residues was found to lead to the production of soluble hyperbranched poly(aryl ether)s. The polymerization process was influenced by the solvents employed, monomer concentration, and the reaction time. An increase in the monomer concentration and reaction time was found to lead to an increase in the molecular weight characteristics of the resulting polymers as determined by gel permeation chromatography (GPC). The composition and architecture of the polyethers were determined by nuclear magnetic resonance (NMR) spectroscopic analysis and were found to be highly complex and dependent on the structure of the monomers used. All of the polymers were found to contain ether linkages formed via carbene insertion into O-H bonds, although polymers derived from phenolic carbenes also contained linkages arising from C-alkylation.