Modulation of Lrp action in Escherichia coli by leucine: effects on non-specific binding of Lrp to DNA.
Lrp is a global regulator of metabolism in Escherichia coli that helps cells respond to changes in environmental conditions. The action of Lrp as a transcriptional activator or repressor is sometimes affected by whether the medium contains exogenous leucine. The abundance of Lrp in cells is relatively high (about 15 microM in monomer), and given the relatively high Lrp binding affinity in vitro for specific binding sites (nanomolar apparent dissociation constants), the expectation is that all binding sites will be saturated with Lrp in vivo. Here we consider the fraction of the total Lrp in cells that is free and the fraction that is bound to DNA. Using minicell-producing strains, we measured the distribution of Lrp between cytoplasm and nucleoid in cells grown under different nutritional conditions and in cells in different phases of growth. In E. coli cells grown in minimal medium to mid-log phase, the ratio of free to DNA-bound Lrp was about 0.67. This ratio decreased about threefold when the cells were grown in minimal medium supplemented with leucine. Our results also confirmed the previous finding that growth rate regulates lrp expression by as much as three to fourfold. Growth rate-regulated lrp expression, along with changes in the extent of non-specific binding, influences the level of free Lrp in vivo over a 16-fold range. We propose that the net effect of these processes is to regulate the relative concentrations of free Lrp hexadecamer and leucine-bound octamer, leading to promoter selection in response to environmental conditions.
Chen
S
S
Department of Molecular Biology and Genetics, Cornell University, Ithaca, NY 14853, USA.
Hao
Z
Z
Bieniek
E
E
Calvo
J M
JM
D003593:Cytoplasm / Q000187:drug effects / Q000378:metabolism; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D004926:Escherichia coli / Q000166:cytology / Q000187:drug effects* / Q000254:growth & development / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D007930:Leucine / Q000494:pharmacology*; D051756:Leucine-Responsive Regulatory Protein; D008957:Models, Genetic; D009876:Operon / Q000235:genetics; D011485:Protein Binding / Q000187:drug effects; D020836:Protein Structure, Quaternary; D021381:Protein Transport / Q000187:drug effects; D013379:Substrate Specificity; D014157:Transcription Factors
D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C497278:Lrp protein, E coli; D014157:Transcription Factors; D051756:Leucine-Responsive Regulatory Protein; D007930:Leucine