Escherichia coli

About: Escherichia coli is a research topic. Over the lifetime, 59041 publications have been published within this topic receiving 2050337 citations. The topic is also known as: E. coli & E coli jdj.

Elongation factor Tu and E1 β subunit of pyruvate dehydrogenase complex act as fibronectin binding proteins in Mycoplasma pneumoniae

Abstract: Summary The interactions between pathogenic bacteria and extracellular matrix (ECM) components markedly influence the initiation and establishment of infection. We have identified two surface proteins of virulent Mycoplasma pneumoniae with molecular masses of 45 and 30 kDa that bind to the ECM constituent, fibronectin (Fn). These Fn-binding proteins (FnBPs) were purified to near homogeneity using Fn-coupled Sepharose 4B-affinity column chromatography, and amino acid sequence analysis of the 45 and the 30 kDa proteins identified them as elongation factor Tu (EF-Tu) and pyruvate dehydrogenase E1 β subunit (PDH-B) respectively. The genes for EF-Tu and PDH-B were cloned, and the entire EF-Tu gene and NH2-terminus of PDH-B (NPDH (pyruvate dehydrogenase E1 β subunit from amino acid 1–244)-B) gene were overexpressed in Escherichia coli. The recombinant proteins, rEF-Tu and rNPDH-B, were purified to homogeneity by His-tag affinity column chromatography and used to immunize rabbits. Purified rEF-Tu and rNPDH-B bound to Fn using a ligand immunoblot assay and ELISA. Immunogold electron microscopy with polyclonal antibodies reactive against rEF-Tu (antirEF-Tu) and rNPDH-B (antirNPDH-B) and whole cell radioimmunoprecipitation (WCRIP) revealed the surface location of these proteins. Adherence of viable M. pneumoniae to immobilized Fn was inhibited by antirEF-Tu and antirNPDH-B antisera in a dose-dependent and cumulative manner. These results demonstrate that M. pneumoniae EF-Tu and PDH-B, in addition to their major cytoplasmic biosynthetic and metabolic roles, can be surface translocated, which confers additional important biological functions.

recA protein of Escherichia coli promotes branch migration, a kinetically distinct phase of DNA strand exchange.

Abstract: The recA protein of Escherichia coli promotes the complete exchange of strands between full-length linear duplex and single-stranded circular DNA molecules of bacteriophage phi X-174, converting more than 50% of the single-stranded DNA into heteroduplex replicative form II-like structures. Kinetically, the reaction can be divided into two phases, formation of short heteroduplex regions (D loops) and extension of the D loops via branch migration. recA protein participates directly in both phases. D loops are formed efficiently in the presence of ATP or the nonhydrolyzable ATP analog adenosine 5'-[gamma-thio]triphosphate, whereas D-loop extension requires continuous ATP hydrolysis. Complete strand exchange requires a stoichiometric amount of recA protein and is strongly stimulated by the single-stranded-DNA-binding protein of E. coli.

Improved method for the isolation of thymine-requiring mutants of escherichia coli.

Abstract: Thymine-requiring mutants of Escherichia coli were rarely found before the observation by Okada, Yanagisawa, and Ryan (Z. Vererbungslehre 92:403, 1961) that cultures of bacteria grown in the presence of high concentrations of aminopterin, thymine, purines, and serine contain a surprising number of cells unable to synthesize their own supply of thymine. They showed that these thymine-dependent mutants could grow much faster in the presence of aminopterin than the parent strains, even though both were supplied with adequate amounts of those metabolites whose syntheses are blocked by aminopterin. These observations provided the basis for a widely used method of isolation of this special class of mutants. The greater efficiency of a simplified medium was reported by Okada, Homma, and Sonohara (J. Bacteriol. 84:602, 1962). We also found that the yield of thymutants was greatly increased if a completely minimal medium, usually M9, containing only thymine (50 ,g/ml) in addition to aminopterin (300 ,ug/ml) was used as the isolation medium. The absence of the other metabolites, for whose synthesis tetrahydrofolate is required, appears to increase the selection pressure in favor of the thymine-dependent mutants. In some experiments, after 48 hr of growth in this medium, the percentage of colony-formers unable to make their own supply of thymine approached 100%. In these experiments, it was much more difficult to isolate thymutants from strains that require vitamin B1 (thiamine). Pine (J. Bacteriol. 79:827, 1960) presented evidence that aminopterin enters the cells of Bacillus subtilis mainly by active transport by the thiamine permease, and that the addition of small amounts of thiamine or a similar pyrimidine affords a great deal of protection against the growth inhibition caused by aminopterin (Pine, J. Bacteriol. 79:835, 1960). It is likely that the added B1 in the medium similarly protects the B1-requiring strains of E. coli. The high concentrations of aminopterin required to inhibit E. coli are necessary because, like B.

DNA Sequence of a ColV Plasmid and Prevalence of Selected Plasmid-Encoded Virulence Genes among Avian Escherichia coli Strains

Abstract: ColV plasmids have long been associated with the virulence of Escherichia coli, despite the fact that their namesake trait, ColV production, does not appear to contribute to virulence. Such plasmids or their associated sequences appear to be quite common among avian pathogenic E. coli (APEC) and are strongly linked to the virulence of these organisms. In the present study, a 180-kb ColV plasmid was sequenced and analyzed. This plasmid, pAPEC-O2-ColV, possesses a 93-kb region containing several putative virulence traits, including iss, tsh, and four putative iron acquisition and transport systems. The iron acquisition and transport systems include those encoding aerobactin and salmochelin, the sit ABC iron transport system, and a putative iron transport system novel to APEC, eit. In order to determine the prevalence of the virulence-associated genes within this region among avian E. coli strains, 595 APEC and 199 avian commensal E. coli isolates were examined for genes of this region using PCR. Results indicate that genes contained within a portion of this putative virulence region are highly conserved among APEC and that the genes of this region occur significantly more often in APEC than in avian commensal E. coli. The region of pAPEC-O2-ColV containing genes that are highly prevalent among APEC appears to be a distinguishing trait of APEC strains.

Plasmid Replicon Typing of Commensal and Pathogenic Escherichia coli Isolates

Abstract: Despite the critical role of plasmids in horizontal gene transfer, few studies have characterized plasmid relatedness among different bacterial populations. Recently, a multiplex PCR replicon typing protocol was developed for classification of plasmids occurring in members of the Enterobacteriaceae. Here, a simplified version of this replicon typing procedure which requires only three multiplex panels to identify 18 plasmid replicons is described. This method was used to screen 1,015 Escherichia coli isolates of avian, human, and poultry meat origin for plasmid replicon types. Additionally, the isolates were assessed for their content of several colicin-associated genes. Overall, a high degree of plasmid variability was observed, with 221 different profiles occurring among the 1,015 isolates examined. IncFIB plasmids were the most common type identified, regardless of the source type of E. coli. IncFIB plasmids occurred significantly more often in avian pathogenic E. coli (APEC) and retail poultry E. coli (RPEC) than in uropathogenic E. coli (UPEC) and avian and human fecal commensal E. coli isolates (AFEC and HFEC, respectively). APEC and RPEC were also significantly more likely than UPEC, HFEC, and AFEC to possess the colicin-associated genes cvaC, cbi, and/or cma in conjunction with one or more plasmid replicons. The results suggest that E. coli isolates contaminating retail poultry are notably similar to APEC with regard to plasmid profiles, with both generally containing multiple plasmid replicon types in conjunction with colicin-related genes. In contrast, UPEC and human and avian commensal E. coli isolates generally lack the plasmid replicons and colicin-related genes seen in APEC and RPEC, suggesting limited dissemination of such plasmids among these bacterial populations.