Deoxyribonucleic acid (DNA)-dependent ribonucleic acid (RNA) polymerase (EC 22.214.171.124) isolated from a rifampin-sensitive strain of Mycobacterium smegmatis was 90% inhibited by 1 μg of rifampin per ml; enzyme from a rifampin-resistant mutant was not affected by this concentration of antibiotic. Inhibition of phenylalanine-1-14C incorporation by rifampin in growing cultures was complete about 6 min after addition of antibiotic. Under the same conditions, uracil-2-14c incorporated was blocked after 1.5 to 2 min. Rifampin kills M. smegmatis very slowly. When rifampin-inhibited cultures were transferred to a rifampin-free medium, there was a partial resumption of uracil-2-14C incorporation, even in the presence of chloramphenicol. We conclude that a primary event in the inhibition of M. smegmatis by rifampin is the block of DNA-dependent RNA polymerase
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