Biomaterial Database

Interactions of Escherichia coli transcription termination factor rho with RNA. I. Binding stoichiometries and free energies. 1988

J A McSwiggen, and D G Bear, and P H von Hippel

Institute of Molecular Biology, University of Oregon, Eugene 97403.

In this paper we examine the binding of Escherichia coli transcription termination factor rho to single-stranded RNA. Random polyribonucleotide copolymers containing low ratios of the fluorescent base 1,N6-ethenoadenosine have been synthesized using polynucleotide phosphorylase. Binding of rho to these polynucleotides elicits a significant increase in fluorescence, thus allowing either the direct monitoring of the titration of these polynucleotides with rho or measurement of the competitive displacement of the protein from these probes with other nucleic acids, even in the presence of biologically significant concentrations of ATP. By these techniques, it is shown that the binding site size (n) of rho protein to polynucleotides is 13(+/- 1) nucleotide residues per rho monomer (or 78(+/- 6) nucleotide residues per rho hexamer). Binding constants (K) and co-operativity parameters (omega) for the binding of rho to these polynucleotides have been measured as a function of nucleotide composition and of salt concentration. The results show that the affinity of rho for cytosine residues is quite strong and salt concentration independent, whilst binding to uridine residues is somewhat weaker and very salt concentration dependent. Poly(rC) and poly(dC) bind to rho competitively and with equal affinity and site size, although poly(rC) is the strongest cofactor for activating rho-dependent ATPase and poly(dC) has no ATPase cofactor activity at all. It is also shown that ATP (or ADP or ATP-gamma-S) binding does not change the binding site size of rho on RNA nor decrease its affinity for RNA binding. Circular dichroism measurements of rho binding to phage R17 RNA suggest that the affinity (K omega) of rho for RNA may be increased by ATP. The possible significance of these results for models of rho-dependent transcription termination is discussed in the companion paper.

UI	MeSH Term	Description	Entries
D007700	Kinetics	The rate dynamics in chemical or physical systems.
D008433	Mathematics	The deductive study of shape, quantity, and dependence. (From McGraw-Hill Dictionary of Scientific and Technical Terms, 6th ed)	Mathematic
D004926	Escherichia coli	A species of gram-negative, facultatively anaerobic, rod-shaped bacteria (GRAM-NEGATIVE FACULTATIVELY ANAEROBIC RODS) commonly found in the lower part of the intestine of warm-blooded animals. It is usually nonpathogenic, but some strains are known to produce DIARRHEA and pyogenic infections. Pathogenic strains (virotypes) are classified by their specific pathogenic mechanisms such as toxins (ENTEROTOXIGENIC ESCHERICHIA COLI), etc.	Alkalescens-Dispar Group,Bacillus coli,Bacterium coli,Bacterium coli commune,Diffusely Adherent Escherichia coli,E coli,EAggEC,Enteroaggregative Escherichia coli,Enterococcus coli,Diffusely Adherent E. coli,Enteroaggregative E. coli,Enteroinvasive E. coli,Enteroinvasive Escherichia coli
D005456	Fluorescent Dyes	Chemicals that emit light after excitation by light. The wave length of the emitted light is usually longer than that of the incident light. Fluorochromes are substances that cause fluorescence in other substances, i.e., dyes used to mark or label other compounds with fluorescent tags.	Flourescent Agent,Fluorescent Dye,Fluorescent Probe,Fluorescent Probes,Fluorochrome,Fluorochromes,Fluorogenic Substrates,Fluorescence Agents,Fluorescent Agents,Fluorogenic Substrate,Agents, Fluorescence,Agents, Fluorescent,Dyes, Fluorescent,Probes, Fluorescent,Substrates, Fluorogenic
D000244	Adenosine Diphosphate	Adenosine 5'-(trihydrogen diphosphate). An adenine nucleotide containing two phosphate groups esterified to the sugar moiety at the 5'-position.	ADP,Adenosine Pyrophosphate,Magnesium ADP,MgADP,Adenosine 5'-Pyrophosphate,5'-Pyrophosphate, Adenosine,ADP, Magnesium,Adenosine 5' Pyrophosphate,Diphosphate, Adenosine,Pyrophosphate, Adenosine
D000255	Adenosine Triphosphate	An adenine nucleotide containing three phosphate groups esterified to the sugar moiety. In addition to its crucial roles in metabolism adenosine triphosphate is a neurotransmitter.	ATP,Adenosine Triphosphate, Calcium Salt,Adenosine Triphosphate, Chromium Salt,Adenosine Triphosphate, Magnesium Salt,Adenosine Triphosphate, Manganese Salt,Adenylpyrophosphate,CaATP,CrATP,Manganese Adenosine Triphosphate,MgATP,MnATP,ATP-MgCl2,Adenosine Triphosphate, Chromium Ammonium Salt,Adenosine Triphosphate, Magnesium Chloride,Atriphos,Chromium Adenosine Triphosphate,Cr(H2O)4 ATP,Magnesium Adenosine Triphosphate,Striadyne,ATP MgCl2
D000495	Allosteric Site	A site on an enzyme which upon binding of a modulator, causes the enzyme to undergo a conformational change that may alter its catalytic or binding properties.	Allosteric Sites,Site, Allosteric,Sites, Allosteric
D001665	Binding Sites	The parts of a macromolecule that directly participate in its specific combination with another molecule.	Combining Site,Binding Site,Combining Sites,Site, Binding,Site, Combining,Sites, Binding,Sites, Combining
D012234	Rho Factor	A protein which effects termination of RNA synthesis during the genetic transcription process by dissociating the ternary transcription complex RNA;-RNA POLYMERASE DNA at the termination of a gene.	E Coli Transcription Termination Factor,Factor, Rho
D012313	RNA	A polynucleotide consisting essentially of chains with a repeating backbone of phosphate and ribose units to which nitrogenous bases are attached. RNA is unique among biological macromolecules in that it can encode genetic information, serve as an abundant structural component of cells, and also possesses catalytic activity. (Rieger et al., Glossary of Genetics: Classical and Molecular, 5th ed)	RNA, Non-Polyadenylated,Ribonucleic Acid,Gene Products, RNA,Non-Polyadenylated RNA,Acid, Ribonucleic,Non Polyadenylated RNA,RNA Gene Products,RNA, Non Polyadenylated