Induction of cadBA in an Escherichia coli lysine auxotroph transformed with a cad-gfp transcriptional fusion
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CadBA functions as a part of overall Escherichia coli response to low extracellular pH. A gfpmut3 structural gene transcriptionally fused to the cadBA promoter (Pcad) was used as a reporter to monitor changes in intracellular lysine as a potential factor influencing cadBA induction. Different patterns of cadBA induction were observed in two E. coli strains with different lysine biosynthetic capabilities. In E. coli ZK126 (pJBA25-Pcad), a lysine prototroph, maximum levels of induction were detected 3 h after the transfer of bacterial cells under inducing conditions (pH 5.8; 3.4 μM extracellular lysine). The induction subsequently decreased until hour 7 after which no further change in expression was observed. However, in the lysine depleted strain E. coli ATCC 23812 (pJBA25-Pcad) which is an auxotroph for lysine, no decrease in cadBA expression was observed over time under the same induction conditions. Although no time dependent statistical differences in intracellular lysine were observed, bacterial cells depleted for no longer than 4 h (1.38 ± 0.25 μmol lysine/g cell dry weight) exhibited more rapid induction of cadBA (after 3 h) and a lower maximum level of induction compared to cells with relatively lower intracellular lysine (approximately 1.08 μmol/g cell dry weight). For the latter, the detectable level of induction was delayed for 1 h but the maximum level of induction response was higher.
KeywordsEscherichia coli Lysine auxotroph cad operon Green fluorescent protein Transcriptional fusion Intracellular lysine
This research was supported by Hatch grant H8311 administered by the Texas Agricultural Experiment Station and Texas Advanced Technology Program, grant #: 000517-0220-2001.The authors would like to express their thanks to Dr. Deborah Siegele (Texas A&M University, College Station, TX) and Julia Sonka (University of Arkansas, Fayetteville, AR) for assistance in the preparation of this manuscript.
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