A resonance Raman study on the interaction of rifampicin with Escherichia coli RNA polymerase.
The technique of resonance Raman spectroscopy has been used to investigate the interaction of the antibiotic rifampicin with Escherichia coli RNA polymerase. Spectra were analyzed by generating the first derivative of each recorded spectrum using the Savitsky-Golay algorithm. The only band that shifted significantly in the resonance Raman spectrum of rifampicin upon the formation of the drug-core polymerase complex was the amide III band. It underwent an 8 cm-1 shift from 1306 cm-1 in aqueous solution to 1314 cm-1. A comparable shift was observed for the rifampicin-holoenzyme complex. Thus, the interaction of the sigma subunit with the core polymerase does not significantly alter the manner in which rifampicin interacts with RNA polymerase. The nature of this shift has been analyzed further by recording the resonance Raman spectrum of rifampicin in a variety of solvents with different hydrogen-bonding solvents (benzene and carbon disulfide) the amide III band was observed at approximately 1220 cm-1; in dimethyl sulfoxide, a weak hydrogen-bond acceptor, 1274 cm-1; in water, a strong hydrogen-bonding solvent, 1306 cm-1; and finally, in triethylamine, a stronger hydrogen-bonding solvent than water, it was observed at 1314 cm-1. Thus, as the hydrogen-bonding ability of the solvent increased, the amide III band shifted to higher frequency. Based on these results, the rifampicin binding site in RNA polymerase provides a stronger hydrogen-bonding environment for the amidic proton of rifampicin than is encountered when rifampicin is free in aqueous solution.
D001665:Binding Sites; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology*; D006860:Hydrogen Bonding; D012293:Rifampin / Q000378:metabolism*; D012997:Solvents; D013059:Spectrum Analysis, Raman
D012997:Solvents; D012321:DNA-Directed RNA Polymerases; D012293:Rifampin