scholarly journals Kinetic Properties of Four Plasmid-Mediated AmpC β-Lactamases

2005 ◽  
Vol 49 (10) ◽  
pp. 4240-4246 ◽  
Author(s):  
Cédric Bauvois ◽  
Akiko Shimizu Ibuka ◽  
Almeida Celso ◽  
Jimena Alba ◽  
Yoshikazu Ishii ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Kinetic Parameters ◽  
Kinetic Properties ◽  
Kinetic Characterization ◽  
Heterologous Production ◽  
Periplasmic Space ◽  
E Coli ◽  
Class C ◽  
Susceptibility Patterns

ABSTRACT The heterologous production in Escherichia coli, the purification, and the kinetic characterization of four plasmid-encoded class C β-lactamases (ACT-1, MIR-1, CMY-2, and CMY-1) were performed. Except for their instability, these enzymes are very similar to the known chromosomally encoded AmpC β-lactamases. Their kinetic parameters did not show major differences from those obtained for the corresponding chromosomal enzymes. However, the Km values of CMY-2 for cefuroxime, cefotaxime, and oxacillin were significantly decreased compared to those of the chromosomal AmpC enzymes. Finally, the susceptibility patterns of different E. coli hosts producing a plasmid- or a chromosome-encoded class C enzyme toward β-lactam antibiotics are mainly due to the overproduction of the β-lactamase in the periplasmic space of the bacteria rather than to a specific catalytic profile of the plasmid-encoded β-lactamases.

scholarly journals Characterization of the New AmpC β-Lactamase FOX-8 Reveals a Single Mutation, Phe313Leu, Located in the R2 Loop That Affects Ceftazidime Hydrolysis

2013 ◽  
Vol 57 (10) ◽  
pp. 5158-5161 ◽  
Author(s):  
Francisco José Pérez-Llarena ◽  
Frédéric Kerff ◽  
Laura Zamorano ◽  
María Carmen Fernández ◽  
Maria Luz Nuñez ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Kinetic Analysis ◽  
Clinical Strain ◽  
Single Mutation ◽  
Content Type ◽  
E Coli ◽  
Fold Reduction ◽  
Class C

ABSTRACTA novel class C β-lactamase (FOX-8) was isolated from a clinical strain ofEscherichia coli. The FOX-8 enzyme possessed a unique substitution (Phe313Leu) compared to FOX-3. IsogenicE. colistrains carrying FOX-8 showed an 8-fold reduction in resistance to ceftazidime relative to FOX-3. In a kinetic analysis, FOX-8 displayed a 33-fold reduction inkcat/Kmfor ceftazidime compared to FOX-3. In the FOX family of β-lactamases, the Phe313 residue located in the R2 loop affects ceftazidime hydrolysis and alters the phenotype ofE. colistrains carrying this variant.

scholarly journals Characterization of proteobacterial plasmid integron-encoded qac efflux pump sequence diversity and quaternary ammonium compound antiseptic selection in E. coli grown planktonically and as biofilms

2021 ◽  
Author(s):  
Carmine J. Slipski ◽  
Taylor R. Jamieson-Datzkiw ◽  
George G. Zhanel ◽  
Denice C. Bay
Keyword(s):  
Escherichia Coli ◽  
Quaternary Ammonium ◽  
Susceptibility Testing ◽  
Efflux Pump ◽  
Multidrug Resistant ◽  
Quaternary Ammonium Compound ◽  
Ammonium Compound ◽  
E Coli ◽  
Susceptibility Patterns

Qac efflux pumps from proteobacterial multidrug-resistant plasmids are integron-encoded and confer resistance to quaternary ammonium compound (QAC) antiseptics, however, many are uncharacterized and misannotated. A survey of >2000 plasmid-encoded qac identified 37 unique qac sequences that correspond to one of five representative motifs: QacE, QacEΔ1, QacF/L, QacH/I, and QacG. Antimicrobial susceptibility testing of each cloned qac member in Escherichia coli , highlighted distinctive antiseptic susceptibility patterns that were most prominent when cells grew as biofilms.

Purification and characterization of D-β-hydroxybutyrate dehydrogenase expressed in Escherichia coli

1993 ◽  
Vol 71 (7-8) ◽  
pp. 406-410
Author(s):  
Les Jones ◽  
Sharon Churchill ◽  
Perry Churchill
Keyword(s):  
Escherichia Coli ◽  
Kinetic Parameters ◽  
Functional Properties ◽  
Rat Liver ◽  
Specific Activity ◽  
Purification And Characterization ◽  
E Coli ◽  
Specific Activities ◽  
Viable Method

D-β-Hydroxybutyrate dehydrogenase (BDH), a lipid-requiring enzyme, has been cloned into pUC18, expressed in Escherichia coli, and purified to homogeneity. The apoenzyme, i.e., the enzyme devoid of phospholipid, has no activity, but can be activated by phospholipid to a specific activity of 129 μmol/(min∙mg). The functional properties of the enzyme expressed in E. coli were compared with the enzyme purified from rat liver. The specific activities, kinetic parameters, and phospholipid activation profiles were virtually identical. These results indicate that the expression of the enzyme in E. coli is a viable method for producing active functional BDH and should allow for the production of specifically altered BDH molecules.Key words: D-β-hydroxybutyrate dehydrogenase, cloning, expression, lipid requiring.

scholarly journals Escherichia coli Malic Enzymes: Two Isoforms with Substantial Differences in Kinetic Properties, Metabolic Regulation, and Structure

2007 ◽  
Vol 189 (16) ◽  
pp. 5937-5946 ◽  
Author(s):  
Federico P. Bologna ◽  
Carlos S. Andreo ◽  
María F. Drincovich
Keyword(s):  
Escherichia Coli ◽  
Metal Ion ◽  
Metabolic Regulation ◽  
Oxidative Decarboxylation ◽  
Kinetic Properties ◽  
Oligomeric State ◽  
E Coli ◽  
Divalent Metal Ion ◽  
High Degree

ABSTRACT Malic enzymes (MEs) catalyze the oxidative decarboxylation of malate in the presence of a divalent metal ion. In eukaryotes, well-conserved cytoplasmic, mitochondrial, and plastidic MEs have been characterized. On the other hand, distinct groups can be detected among prokaryotic MEs, which are more diverse in structure and less well characterized than their eukaryotic counterparts. In Escherichia coli, two genes with a high degree of homology to ME can be detected: sfcA and maeB. MaeB possesses a multimodular structure: the N-terminal extension shows homology to ME, while the C-terminal extension shows homology to phosphotransacetylases (PTAs). In the present work, a detailed characterization of the products of E. coli sfcA and maeB was performed. The results indicate that the two MEs exhibit relevant kinetic, regulatory, and structural differences. SfcA is a NAD(P) ME, while MaeB is a NADP-specific ME highly regulated by key metabolites. Characterization of truncated versions of MaeB indicated that the PTA domain is not essential for the ME reaction. Nevertheless, truncated MaeB without the PTA domain loses most of its metabolic ME modulation and its native oligomeric state. Thus, the association of the two structural domains in MaeB seems to facilitate metabolic control of the enzyme. Although the PTA domain in MaeB is highly similar to the domains of proteins with PTA activity, MaeB and its PTA domain do not exhibit PTA activity. Determination of the distinct properties of recombinant products of sfcA and maeB performed in the present work will help to clarify the roles of MEs in prokaryotic metabolism.

scholarly journals Cloning and Characterization of a Chromosomal Class C β-Lactamase and Its Regulatory Gene in Laribacter hongkongensis

2005 ◽  
Vol 49 (5) ◽  
pp. 1957-1964 ◽  
Author(s):  
Susanna K. P. Lau ◽  
Pak-leung Ho ◽  
Maria W. S. Li ◽  
Hoi-wah Tsoi ◽  
Raymond W. H. Yung ◽  
...  
Keyword(s):  
Genomic Dna ◽  
Southern Hybridization ◽  
Regulatory Gene ◽  
Kinetic Properties ◽  
Restriction Fragments ◽  
Regulatory Systems ◽  
Laribacter Hongkongensis ◽  
Consensus Motifs ◽  
Class C

ABSTRACT Laribacter hongkongensis, a newly discovered bacterium recently shown to be associated with community-acquired gastroenteritis, is generally resistant to most β-lactams except the carbapenems. We describe the cloning and characterization of a novel chromosomal class C β-lactamase and its regulatory gene in L. hongkongensis. Two genes, ampC and ampR, were cloned by inserting restriction fragments of genomic DNA from L. hongkongensis strain HLHK5 into pBK-CMV to give the recombinant plasmid pBK-LHK-5. The ampR and ampC genes and their promoters were divergently oriented, with the ampR gene immediately upstream of the ampC gene and an intercistronic Lys-R motif, typical of inducible ampC-ampR regulatory systems. The deduced amino acid sequence of the cloned AmpC β-lactamase (pI 8.1) contained consensus motifs characteristic of class C β-lactamases but had identities no greater than 46% to known class C β-lactamases. The kinetic properties of this AmpC were also compatible with those of a class C β-lactamase. PCR of 20 clinical isolates of L. hongkongensis, including HLHK5, showed the presence of both ampC and ampR genes in all isolates. Southern hybridization suggested that the ampC gene of HLHK5 was chromosomally encoded. Subcloning experiments showed that the expression of the ampC gene of HLHK5 was regulated by its ampR gene, which acts as a repressor. The β-lactamase characterized from strain HLHK5 was named LHK-5 (gene, bla LHK-5) and represents the first example of AmpC β-lactamase in the β subdivision of proteobacteria.

scholarly journals Functional expression of the eukaryotic proton pump rhodopsin OmR2 in Escherichia coli and its photochemical characterization

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Masuzu Kikuchi ◽  
Keiichi Kojima ◽  
Shin Nakao ◽  
Susumu Yoshizawa ◽  
Shiho Kawanishi ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Proton Pump ◽  
Expression System ◽  
Spectroscopic Characterization ◽  
Functional Expression ◽  
Proton Pumping ◽  
E Coli ◽  
Oxyrrhis Marina ◽  
Domains Of Life

AbstractMicrobial rhodopsins are photoswitchable seven-transmembrane proteins that are widely distributed in three domains of life, archaea, bacteria and eukarya. Rhodopsins allow the transport of protons outwardly across the membrane and are indispensable for light-energy conversion in microorganisms. Archaeal and bacterial proton pump rhodopsins have been characterized using an Escherichia coli expression system because that enables the rapid production of large amounts of recombinant proteins, whereas no success has been reported for eukaryotic rhodopsins. Here, we report a phylogenetically distinct eukaryotic rhodopsin from the dinoflagellate Oxyrrhis marina (O. marina rhodopsin-2, OmR2) that can be expressed in E. coli cells. E. coli cells harboring the OmR2 gene showed an outward proton-pumping activity, indicating its functional expression. Spectroscopic characterization of the purified OmR2 protein revealed several features as follows: (1) an absorption maximum at 533 nm with all-trans retinal chromophore, (2) the possession of the deprotonated counterion (pKa = 3.0) of the protonated Schiff base and (3) a rapid photocycle through several distinct photointermediates. Those features are similar to those of known eukaryotic proton pump rhodopsins. Our successful characterization of OmR2 expressed in E. coli cells could build a basis for understanding and utilizing eukaryotic rhodopsins.

scholarly journals Isolation and characterization of escherichia coli in ready-to-eat foods vended in Islamic University, Kushtia

1970 ◽  
Vol 18 ◽  
pp. 99-103 ◽  
Author(s):  
S Biswas ◽  
MAK Parvez ◽  
M Shafiquzzaman ◽  
S Nahar ◽  
MN Rahman
Keyword(s):  
Escherichia Coli ◽  
Pattern Analysis ◽  
University Campus ◽  
Rflp Pattern ◽  
E Coli ◽  
Fish Meat ◽  
Isolation And Characterization ◽  
Food Borne ◽  
Identification And Characterization

Context: Escherichia coli is shed in the feces of warm blooded animals and humans and thus potential for public health. Detection and characterization of E. coli in the ready-to-eat (RTE) foods concerns due to their presence indicates fecal contamination of the food.   Objective: To identify, characterize and RFLP pattern analysis of E. coli isolated from RTE foods vended in Islamic University campus, Kushtia.   Materials and Methods: Fifty samples from four types of consumed foods in six student halls of residence, some temporary restaurants of Islamic University, Kushtia were assessed for bacterial contamination by standard methods. Identification and characterization of E. coli isolates were performed using IMViC tests. Genomic DNA was used to perform RFLP pattern analysis.   Results: Thirty seven out of 50 (74%) examined samples of RTE foods had E. coli contamination. The highest number of E. coli was isolated from vegetable oriented RTE foods (90.90%) and fish, meat and cereals samples were also significantly E. coli positive. RFLP profiling of two E. coli isolates were observed.   Conclusion: The results of this study provide evidence that some RTE foods had unsatisfactory levels of contamination with E. coli. Thus street vended RTE food could be important potential vehicles for food-borne diseases. Molecular characterization may be exploited to identify food borne pathogen among different species.  Keywords: Ready-to-eat foods; Escherichia coli; RFLP pattern DOI: http://dx.doi.org/10.3329/jbs.v18i0.8783 JBS 2010; 18(0): 99-103

scholarly journals Evaluation of antipyretic activity of hydroalcoholic extract of Corchorus depressus Linn. in Escherichia coli–induced pyretic rabbits

2018 ◽  
Vol 16 ◽  
pp. 205873921879295
Author(s):  
Saeed Ahmad ◽  
Muhammad Akram ◽  
Syed Muhammad Ali Shah ◽  
Sabira Sultana
Keyword(s):  
Escherichia Coli ◽  
Positive Control ◽  
Negative Control ◽  
The Body ◽  
Hydroalcoholic Extract ◽  
Antipyretic Effect ◽  
E Coli ◽  
Antipyretic Activity ◽  
Experimental Treatments

This study was conducted to investigate the antipyretic effect of the hydroalcoholic extract of Corchorus depressus Linn. against Escherichia coli ( E. coli)-induced pyrexia in rabbits. Hydroalcohalic extracts of C. depressus were given orally at 25, 50, and 100 mg/kg for antipyretic affect in E. coli-induced fever in rabbits. The animals were divided into five groups of five each. Among these five groups, three received various doses of experimental treatments, whereas the fourth one served as positive control and received paracetamol. The fifth group of animals served as negative control and received no treatment. The body temperature of the rabbits was measured rectally over a period of 5 h. C. depressus exhibited better effects at dose rate of 25, 50, and 100 mg/kg. The hydroalcoholic extract of C. depressus has significant antipyretic effect. These results lend support to the popular use of C. depressus in traditional medicine as a remedy for pyrexia and suggest that the characterization of the principles for such activity deserves further investigation.

Characterization of novel ybjG and dacC variants in Escherichia coli

2013 ◽  
Vol 62 (11) ◽  
pp. 1728-1734 ◽  
Author(s):  
Dongguo Wang ◽  
Enping Hu ◽  
Jiayu Chen ◽  
Xiulin Tao ◽  
Katelyn Gutierrez ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Multiple Drug Resistance ◽  
Multiple Drug ◽  
The Novel ◽  
Conventional Pcr ◽  
E Coli ◽  
Novel Variants ◽  
Restriction Sites ◽  
Genetic Structures

A total of 69 strains of Escherichia coli from patients in the Taizhou Municipal Hospital, China, were isolated, and 11 strains were identified that were resistant to bacitracin, chloramphenicol, tetracycline and erythromycin. These strains were PCR positive for at least two out of three genes, ybjG, dacC and mdfA, by gene mapping with conventional PCR detection. Conjugation experiments demonstrated that these genes existed in plasmids that conferred resistance. Novel ybjG and dacC variants were isolated from E. coli strains EC2163 and EC2347, which were obtained from the sputum of intensive care unit patients. Genetic mapping showed that the genes were located on 8200 kb plasmid regions flanked by EcoRI restriction sites. Three distinct genetic structures were identified among the 11 PCR-positive strains of E. coli, and two contained the novel ybjG and dacC variants. The putative amino acid differences in the ybjG and dacC gene variants were characterized. These results provide evidence for novel variants of ybjG and dacC, and suggest that multiple drug resistance in hospital strains of E. coli depends on the synergistic function of ybjG, dacC and mdfA within three distinct genetic structures in conjugative plasmids.

Sign in / Sign up

Export Citation Format

Share Document