Active groups of bicyclomycin and the reaction with thiols

The binding of [14C]bicyclomycin to whole cells of E. coli and to the inner membrane proteins was inhibited by dithiothreitol and 2-mercaptoethanol. The reactivity of the drug with the sulfhydryl group was further studied, using methanethiol as a model compound. The kinetics revealed that the reaction was of pseudo-first-order in excess of thiolate anion. Analysis with gas chromatography-mass spectrometry showed that the main product was an adduct of thiol with bicyclomycin in an equal molar ratio. The structure of the adduct was determined by 1H-NMR spectrometry, showing that thiolate attacked the olefinic double bond of the antibiotic. 3'-Acyl derivatives of bicyclomycin did not significantly affect the binding of [14C] bicyclomycin to inner membrane proteins of E. coli. The results suggested that 4,5-double bond hydrocarbons and 3'-hydroxy group of bicyclomycin participate in the binding to E. coli inner membrane proteins, which are presumably the receptors of the antibiotic. The olefinic double bond seems to be the active center of bicyclomycin, reacting with the sulfhydryl group of the receptor protein, although the whole molecular is needed for the activity.