Defective ribosomes in chloramphenicol-treated Escherichia coli

Abstract

Escherichia coli Q13 cells were treated with chloramphenicol for varying periods of time. Upon removal of this drug, the cells exhibited a lag period before the resumption of the normal rate of protein synthesis. This lag period was approximately proportional to the time the cells had been exposed to the drug. Treatment with chloramphenicol not only inhibited protein synthesis but also impaired the protein-synthesizing system of the cells. Ribosomes isolated from the cells exposed to the drug for more than 90 minutes lost 12 to 14% of their protein. When the ribosomes were washed with 1 m-NH 4Cl, no difference in protein content between the ribosomes from normal and drug-treated cells was detected. The ribosomes from drug-treated cells showed a marked decrease in bacteriophage MS2 RNA-specific binding of N-formyknethionyl-tRNA and MS2 RNA-directed protein synthesis when compared to normal ribosomes. No decrease in polyuridylic acid-directed polyphenylalanine synthesis was observed. Supernatant protein fraction from drug-treated cells did not appear to be defective. Addition of crude initiation protein factors prepared from normal ribosomes restored the ability of ribosomes from drug-treated cells to initiate and translate MS2 RNA normally. Acrylamide gel electrophoretic analysis indicated that initiation factor F2 was substantially decreased in the ribosomes from ehloramphenicol-treated cells. The protein fractions removed from ribosomes of normal and drug-treated cells by washing with 1 m-NH 4Cl were applied to DEAE-cellulose columns. Initiation factors F2 and F3 were either defective or substantially reduced in ribosomes from drug-treated cells. Addition of F2 and F3 derived from normal ribosomes restored the ability of the ribosomes from drug-treated cells to carry out MS2 RNA-directed protein synthesis.