Free RNA polymerase in Escherichia coli.
The frequencies of transcription initiation of regulated and constitutive genes depend on the concentration of free RNA polymerase holoenzyme [Rf] near their promoters. Although RNA polymerase is largely confined to the nucleoid, it is difficult to determine absolute concentrations of [Rf] at particular locations within the nucleoid structure. However, relative concentrations of free RNA polymerase at different growth rates, [Rf]rel, can be estimated from the activities of constitutive promoters. Previous studies indicated that the rrnB P2 promoter is constitutive and that [Rf]rel in the vicinity of rrnB P2 increases with increasing growth rate. Recently it has become possible to directly visualize Rf in growing Escherichia coli cells. Here we examine some of the important issues relating to gene expression based on these new observations. We conclude that: (i) At a growth rate of 2 doublings/h, there are about 1000 free and 2350 non-specifically DNA-bound RNA polymerase molecules per average cell (12 and 28%, respectively, of 8400 total) which are in rapid equilibrium. (ii) The reversibility of the non-specific binding generates more than 1000 free RNA polymerase molecules every second in the immediate vicinity of the DNA. Of these, most rebind non-specifically to the DNA within a few ms; the frequency of non-specific binding is at least two orders of magnitude greater than specific binding and transcript initiation. (iii) At a given amount of RNA polymerase per cell, [Rf] and the density of non-specifically DNA-bound RNA polymerase molecules along the DNA both vary reciprocally with the amount of DNA in the cell. (iv) At 2 doublings/h an E. coli cell contains, on the average, about 1 non-specifically bound RNA polymerase per 9 kbp of DNA and 1 free RNA polymerase per 20 kbp of DNA. However some DNA regions (i.e. near active rRNA operons) may have significantly higher than average [Rf].
Department of Medical Genetics, University of Wisconsin-Madison, USA. Electronic address: mpatrick@wisc.edu.
Michael
M
Patrick
Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, VA, USA. Electronic address: dennisp@janelia.hhmi.org.
Patrick P
PP
Dennis
Department of Cell and Molecular Biology, BMC, Uppsala University, Box 596, S-751 24 Uppsala, Sweden. Electronic address: ehrenberg@xray.bmc.uu.se.
Mans
M
Ehrenberg
Department of Molecular and Cell Biology, University of Texas at Dallas, USA. Electronic address: bremer3@att.net.
Hans
H
Bremer
D000465:Algorithms; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D003593:Cytoplasm / Q000201:enzymology / Q000378:metabolism; D004269:DNA, Bacterial / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004058:Diffusion; D004789:Enzyme Activation; D004792:Enzyme Precursors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology* / Q000254:growth & development / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D020035:Holoenzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D008954:Models, Biological; D009154:Mutation; D011401:Promoter Regions, Genetic; D021381:Protein Transport; D012995:Solubility; D014158:Transcription, Genetic; D012407:rRNA Operon
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; C066312:DnaA protein, Bacteria; D004792:Enzyme Precursors; D029968:Escherichia coli Proteins; D020035:Holoenzymes; D012321:DNA-Directed RNA Polymerases
Escherichia coli; Free RNA polymerase; Non-specifically bound RNA polymerase