Uptake of 14G-Streptomycin by Bacillus megaterium

Abstract

More than 90% of the radioactivity taken up by B. megaterium growing in medium containing inhibitory concentrations of 14c-streptomycin appeared to be identical with streptomycin as shown by its behavior on an ion-exchange resin and by recrystallization with streptomycin derivatives. Streptomycin was not removed from the organisms by washing in growth medium with or without addition of unlabelled streptomycin or in distilled water. It could be extracted with inorganic acids and trichloroacetic acid but not by some other commonly used extraction procedures. When the organisms were converted to protoplasts and these lysed and fractionated up to 99% of the streptomycin in the organisms was recovered in the material sedimented from the cytoplasmic fraction by centrifugation at 105,000 g. Very small amounts of streptomycin were found in the fraction containing protoplast "ghosts", but these may have represented contaminating cytoplasmic material. This distribution may not, however, be a true indication of the location of streptomycin before fractionation since a similar distribution was found when streptomycin was added to lysed protoplasts immediately before fractionation. The uptake of streptomycin was decreased at pH values below 7. Uptake depended on continued synthesis of cell material, and environmental conditions which prevented growth and bactericidal action also considerably decreased uptake of streptomycin. The quantity of streptomycin finally taken up was proportional to the concentration of streptomycin in the growth medium and at the lowest growth-inhibitory concentrations was about 5 x 104 molecules of streptomycin per bacterium.