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. 2002 Oct;184(19):5307-16.
doi: 10.1128/JB.184.19.5307-5316.2002.

"V体育官网" Adaptation of sucrose metabolism in the Escherichia coli wild-type strain EC3132

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Adaptation of sucrose metabolism in the Escherichia coli wild-type strain EC3132

Knut Jahreis et al. J Bacteriol. 2002 Oct.

Abstract

Although Escherichia coli strain EC3132 possesses a chromosomally encoded sucrose metabolic pathway, its growth on low sucrose concentrations (5 mM) is unusually slow, with a doubling time of 20 h. In this report we describe the subcloning and further characterization of the corresponding csc genes and adjacent genes. The csc regulon comprises three genes for a sucrose permease, a fructokinase, and a sucrose hydrolase (genes cscB, cscK, and cscA, respectively). The genes are arranged in two operons and are negatively controlled at the transcriptional level by the repressor CscR. Furthermore, csc gene expression was found to be cyclic AMP-CrpA dependent. A comparison of the genomic sequences of the E. coli strains EC3132, K-12, and O157:H7 in addition to Salmonella enterica serovar Typhimurium LT2 revealed that the csc genes are located in a hot spot region for chromosomal rearrangements in enteric bacteria. The comparison further indicated that the csc genes might have been transferred relatively recently to the E. coli wild-type EC3132 at around the time when the different strains of the enteric bacteria diverged. We found evidence that a mobile genetic element, which used the gene argW for site-specific integration into the chromosome, was probably involved in this horizontal gene transfer and that the csc genes are still in the process of optimal adaptation to the new host VSports手机版. Selection for such adaptational mutants growing faster on low sucrose concentrations gave three different classes of mutants. One class comprised cscR(Con) mutations that expressed all csc genes constitutively. The second class constituted a cscKo operator mutation, which became inducible for csc gene expression at low sucrose concentrations. The third class was found to be a mutation in the sucrose permease that caused an increase in transport activity. .

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"V体育安卓版" Figures

FIG. 1.
FIG. 1.
Detailed comparative map of the csc-dsd regions of the E. coli strains K-12, O157:H7, and EC3132. Genes are represented by boxes, with their orientations indicated by arrows. Regions of homology between E. coli K-12 and the two other strains are shown on a dark grey background; those between O157:H7 and EC3132 are shown on a hatched background. O157:H7-specific DNA is emphasized on a light grey hatched background. The heading numbers refer to the positions of the genes on the E. coli K-12 map (in minutes) and to the relative kilobase coordinates.
FIG. 2.
FIG. 2.
Schematic illustration of the construction of csc hybrid plasmids (A) and nucleotide sequence analysis of the csc promoter-operator region (B). (A) The Csc phenotype for sucrose utilization was tested on MacConkey indicator plates with 1% sucrose (scored as 1+ to 3+, pink to deep-purple colonies). The genes are indicated by boxes, with the gene or promoter orientations shown by arrows. DNA of pJBL101 is represented by dark grey boxes, and DNA of pKJL124 is represented by light grey boxes. The position of mutation in pKJL124 is symbolized by an asterisk. (B) The putative −35 and −10 regions are underlined, as well as the putative ribosome-binding sites (RBS) and the start codons of cscK and cscA. Transcriptional start points (TK) are indicated by letters in boldface italic type, the putative csc operators are boxed, and the putative cAMP-CrpA binding sites are marked with diagonal lines. The mutation of pKJL124 is symbolized by an asterisk.
FIG. 3.
FIG. 3.
Sucrose transport by JM109 harboring cscB expression plasmids. Time courses of active sucrose transport by JM109 expressing wild-type sucrose permease from pTM3132 (triangles) or the Q353H derivative of CscB from pTMB1 (circles) are indicated. Cells were grown aerobically at 37°C until mid-log exponential growth phase, harvested, washed with minimal medium, and immediately tested. The final sucrose concentration in the test was 15 μM. Mean values and standard deviations (error bars) from at least three different experiments are given.

References

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