Characterization of the β-lactamase promoter of pBR322

Russell, Debra; Bennett, George N.

doi:10.1093/nar/9.11.2517

Cited by 48 publications

(24 citation statements)

References 38 publications

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“…Twenty-one of the 23 strains presenting a PL penicillinase phenotype produced the ␤-lactamase TEM-1. Twenty isolates had the TEM-1 type promoter P3 (30), and only 1 had the TEM-2 type promoter pair Pa and Pb (13) characterized by a C323T substitution. One strain produced the ␤-lactamase SHV-1, and one strain produced the ␤-lactamase CARB-2.…”

Section: Resultsmentioning

confidence: 99%

Prevalence of β-Lactamases among 1,072 Clinical Strains of Proteus mirabilis : a 2-Year Survey in a French Hospital

Chanal

Bonnet

Champs

et al. 2000

Antimicrob Agents Chemother

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␤-Lactam resistance was studied in 1,072 consecutive P. mirabilis clinical strains isolated at the ClermontFerrand teaching hospital between April 1996 and March 1998. The frequency of amoxicillin resistance was 48.5%. Among the 520 amoxicillin-resistant isolates, three resistance phenotypes were detected: penicillinase (407 strains [78.3%]), extended-spectrum ␤-lactamase (74 strains [14.2%]), and inhibitor resistance (39 strains [7.5%]). The penicillinase phenotype isolates were divided into three groups according to the level of resistance to ␤-lactams, which was shown to be related to the strength of the promoter. The characterization of the different ␤-lactamases showed that amoxicillin resistance in P. mirabilis was almost always (97%) associated with TEM or TEM-derived ␤-lactamases, most of which evolved via TEM-2.After Escherichia coli, Proteus mirabilis is the most often isolated member of the Enterobacteriaceae in European clinical microbiology laboratories, being isolated more often than Klebsiella pneumoniae (22,26). Wild-type strains of P. mirabilis are susceptible to all penicillins and cephalosporins. However, since 1990, the resistance of the species to ␤-lactams has regularly increased (20,22,26,32). Amoxicillin resistance in P. mirabilis is mainly due to the plasmid-mediated penicillinases TEM-1 and TEM-2. TEM-2 is more frequently encountered in this species than in other Enterobacteriaceae (22, 28); TEM-like penicillinase TEM-57 was recently reported by Bonnet et al. (7).Since 1991, TEM-derived extended-spectrum ␤-lactamases (ESBL) (TEM-3, TEM-8, TEM-10, TEM-21, TEM-24, TEM-26, and TEM-66) in P. mirabilis have been reported (7,11,15,24,27,29; L. Pagani, F. Luzzaro, R. Migliavacca, M. G. Perilli, R. Daturi, G. Lombardi, C. Matti, E. Giacobone, and G. Amicosante, Abstr. 37th Intersci. Conf. Antimicrob. Agents Chemother., abstr. D14, p. 85, 1997). Inhibitor-resistant TEM ␤-lactamases (TEM-44, TEM-65, TEM-73, and TEM-74) in this species have been recently described (7, 9). Finally, non-TEM-derived ␤-lactamases (CMY-3, CMY-4, CEP-1, CTX-M-2, and PER-2) in P. mirabilis have also been reported (4,5,6,8,36). The aim of this 2-year survey was to assess the prevalence of established and newer ␤-lactamases among amoxicillin-resistant P. mirabilis clinical strains isolated from the teaching hospital of Clermont-Ferrand. MATERIALS AND METHODSBacterial strains. All nonduplicate P. mirabilis strains isolated from ClermontFerrand teaching hospital during a 2-year survey (1 April 1996 to 31 March 1998) were included in this study. Isolates were identified by the Rapid ID 32 E system (BioMérieux, La Balme les Grottes, France).Susceptibility testing. Susceptibilities of P. mirabilis isolates were determined by the Rapid ATB E system (BioMérieux). A modified double-disc synergy test (11) was used to detect ESBL. The amoxicillin-resistant isolates were screened for susceptibility to a battery of ␤-lactam antibiotics chosen to facilitate the recognition of the resistance patterns associated with the different ␤-lac...

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Section: Resultsmentioning

confidence: 99%

Prevalence of β-Lactamases among 1,072 Clinical Strains of Proteus mirabilis : a 2-Year Survey in a French Hospital

Chanal

Bonnet

Champs

et al. 2000

Antimicrob Agents Chemother

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“…A pBR322-derived vector, pBREP, was constructed to functionally express the mature form of wild-type rice EPSPS (OsEPSPS; Xu et al, 2002) in E. coli under the control of the bla promoter (Russell and Bennett, 1981). Random mutagenesis via error-prone PCR must be performed on sequences less than 1 kb (Cadwell and Joyce, 1994), thus the OsEPSPS coding sequence was separated into two regions (36-542 bp, 542-1,335 bp) to perform error-prone PCR.…”

Section: Identification Of Osepsps Mutant Conferring Glyphosate Resismentioning

confidence: 99%

Identification of a Glyphosate-Resistant Mutant of Rice 5-Enolpyruvylshikimate 3-Phosphate Synthase Using a Directed Evolution Strategy

Zhou

Wei

et al. 2005

Plant Physiology

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5-Enolpyruvylshikimate 3-phosphate synthase (EPSPS) is a key enzyme in the shikimate pathway and is targeted by the widespectrum herbicide glyphosate. Here, we describe the use of a selection system based on directed evolution to select glyphosate-resistant mutants of EPSPS. Using this system, the rice (Oryza sativa) EPSPS gene, mutagenized by Error-Prone polymerase chain reaction, was introduced into an EPSPS-deficient Escherichia coli strain, AB2829, and transformants were selected on minimal medium by functional complementation. Three mutants with high glyphosate resistance were identified in three independent glyphosate selection experiments. Each mutant contained a C 317 /T transition within the EPSPS coding sequence, causing a change of proline-106 to leucine (P106L) in the protein sequence. Glyphosate resistance assays indicated a 3-fold increase in glyphosate resistance of E. coli expressing the P106L mutant. Affinity of the P106L mutant for glyphosate and phosphoenolpyruvate was decreased about 70-fold and 4.6-fold, respectively, compared to wild-type EPSPS. Analysis based on a kinetic model demonstrates that the P106L mutant has a high glyphosate resistance while retaining relatively high catalytic efficiency at low phosphoenolpyruvate concentrations. A mathematical model derived from the Michaelis-Menten equation was used to characterize the effect of expression level and selection conditions on kinetic (K i and K m ) variation of the mutants. This prediction suggests that the expression level is an important aspect of the selection system. Furthermore, glyphosate resistance of the P106L mutant was confirmed in transgenic tobacco (Nicotiana tabacum), demonstrating the potential for using the P106L mutant in transgenic crops.

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“…a Nucieotide position in the Tn3 sequence (16 (4,8,29). The nucleotide sequence differences that we established between Tn2 and Tn3 are shown in Table 1.…”

Section: Hicisf-hoihaimentioning

confidence: 99%

“…(Coordinates established for the nucleotide sequence of Tn3 [16] are used to identify corresponding regions in other Tn3-like transposons.) It was concluded that this mutant transposon, designated Tn2661, carries two overlapping promoters (Pa and Pb) which result in the initiation of two new transcripts, each at a greater efficiency than that from P3, the natural ,B-lactamase promoter used in Tn3, Tn2660, and pBR322 (4,8,29,33 [28] [ Fig. 1]).…”

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confidence: 99%

Variations between the nucleotide sequences of Tn1, Tn2, and Tn3 and expression of beta-lactamase in Pseudomonas aeruginosa and Escherichia coli

Chen¹,

Clowes²

1987

J Bacteriol

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Using SI nuclease assays, we located the sites of initiation of transcription of the j3-lactamase gene on Tnl and Tn2. Transcription in Tn2, like that in Tn3, occurred from the P3 promoter, whereas transcription in Tnl was initiated by two stronger and overlapping promoters, Pa and Pb. The nucleotide sequences of Tnl and Tn2 were determined over a 1,195-base-pair segment constituting most of the sequences of the tnpR and bla genes and the intervening region. There were six base-pair differences between Tnl and Tn3. One in the bla regulatory region accounted for the presence of the Pa and Pb promoters, and another in the bla structural gene is consistent with the isoelectric focusing difference found between the Tnl and Tn3 enzymes. In contrast, there were 24 base-pair differences between Tn2 and Tn3, most of them clustered in one segment of the tnpR gene.A number of transposons which determine TEM-type P-lactamases were originally found by heteroduplex studies (17) to be closely related. They were later distinguished as Tnl, Tn2, Tn3, etc., because of their derivation from different plasmids (6). Since that time, a number of restriction site similarities and differences among them have been identified (5, 28), but since nucleotide sequence determinations have been published only for Tn3, comparisons at the nucleotide level have not been made. The plasmid R6K (18) specifies streptomycin and ampicillin resistance, the latter due to a ,-lactamase (bla) gene, which determines the prototype of the TEM group of ,-lactamases (12), later defined as TEM-1 (20), and which is carried on the transposon Tn2660 (9). Tn2660 is identical to Tn3 in a number of genetic and physiological properties (9, 10, 32) and in its nucleotide sequence over approximately 500 base pairs (bp) at each of the terminal regions (16, 32) and over 500 bp within the noncoding and the amino-terminal regions of the ,-lactamase gene (8).We have recently shown (8) that a mutation in Tn2660 which gives rise to an approximate 10-fold increase in ,-lactamase (bla) resistance is due to a single base-pair substitution at nucleotide 3773, 177 bases upstream of the bla initiation codon (Fig. 1) Tn3, Tn2660, and pBR322 (4, 8, 29, 33 (Fig. 1), was then isolated and used as a source for some of the segments for nucleotide sequence determinations and also for cloning in the vector pRO1614 to produce pRC401. pRO1614 was constructed from pBR322 (26) by inserting into the PstI site a 1.85-kilobase segment derived from a cryptic P. aeruginosa plasmid that permits replication in P. aeruginosa or E. coli hosts. The 1.85-kilobase segment carries the 1-lactamase region downstream from the PstI locus, so that in pRO1614 the P-lactamase gene is reconstituted and is functional, having retained the BamHI-PstI segment of pBR322, i.e., nucleotides 3740 to 4495 of Tn3. The derived plasmid pRC401, in which the 924-bp BamHI-PstI segment from Tnl is substituted for the corresponding segment of pBR322, was used to observe ,B-lactamase expression in P. aeruginosa or E. coli hosts.T...

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Characterization of the β-lactamase promoter of pBR322

Cited by 48 publications

References 38 publications

Prevalence of β-Lactamases among 1,072 Clinical Strains of Proteus mirabilis : a 2-Year Survey in a French Hospital

Prevalence of β-Lactamases among 1,072 Clinical Strains of Proteus mirabilis : a 2-Year Survey in a French Hospital

Identification of a Glyphosate-Resistant Mutant of Rice 5-Enolpyruvylshikimate 3-Phosphate Synthase Using a Directed Evolution Strategy

Variations between the nucleotide sequences of Tn1, Tn2, and Tn3 and expression of beta-lactamase in Pseudomonas aeruginosa and Escherichia coli

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