Protein Synthesis in Mycobacterium tuberculosis H37Rv and the Effect of Streptomycin in Streptomycin-Susceptible and -Resistant Strains

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Abstract
An efficient in vitro amino acid-incorporating system from Mycobacterium tuberculosis H37Rv was standardized. Ribonucleic acid (RNA) isolated from phage-infected M. smegmatis cells served as natural messenger RNA and directed the incorporation of 14 C-amino acids into protein. The effects of various antitubercular drugs and “known inhibitors” of protein synthesis on amino acid incorporation were studied. Antibiotics like chloramphenicol and tetracycline inhibited mycobacterial protein synthesis, though they failed to prevent the growth of the organism. This failure was shown to be due to the impermeability of mycobacteria to these drugs by use of “membrane-active” agents along with the antibiotics in growth inhibition studies. Several independent streptomycin-resistant mutants of M. tuberculosis H37Rv were isolated. Streptomycin inhibited the incorporation of 14 C-amino acids into proteins by whole cells of a streptomycin-susceptible strain by more than 90%, whereas very little or no inhibition was observed in either high-level or low-level streptomycin-resistant strains. In vitro, streptomycin was an effective inhibitor of susceptible strains, whereas in streptomycin-resistant strains the concentration of streptomycin at which half-maximal inhibition was produced varied according to the resistance of whole cells, and there was a correlation between the two. In one low-level streptomycin-resistant mutant, the in vitro amino acid-incorporating system was as sensitive to various concentrations of streptomycin as the parental type, and a possible involvement of a membrane site in the development of low-level resistance was indicated. Streptomycin susceptibility and high-level resistance were shown to be ribosomal in nature.