Gray W J, Midgley J E
Biochem J. 1971 Apr;122(2):161-9. doi: 10.1042/bj1220161.
A study was made of the kinetics of labelling of the stable ribonucleic acids (rRNA+tRNA) and the unstable mRNA fraction in cultures of Escherichia coli M.R.E.600, inhibited by the addition of 0.1g of rifampicin/l. Labelling was carried out by adding either [2-(14)C]- or [5-(3)H]-uracil as an exogenous precursor of the cellular nucleic acids. From studies using DNA RNA hybridization, the kinetics of the synthesis and degradation of mRNA was followed in the inhibited cultures. Although a considerable proportion of the mRNA labelled in the presence of rifampicin decayed to non-hybridizable products, about 25% was stabilized beyond the point where protein synthesis had finally ceased. It therefore seems unwise to extrapolate the results of studies on mRNA stability in rifampicin-inhibited cultures to the situation existing in the rate of steady growth, where there appears to be little, if any, stable messenger. The kinetics of labelling of RNA in inhibited cultures indicated that the clapsed time from the addition of rifampicin to the point at which radioactivity no longer enters the total cellular ribonucleic acids is a measure of the time required to polymerize a molecule of rRNA. At 37 degrees C, in culture grown in broth, glucose-salts or lactate salts media, exogenous [2-(14)C]uracil entered rifampicin-inhibited cells and was incorporated into RNA for 2 3min after the antibiotic was added. Taking this time as that required to polymerize a complete chain of 23S rRNA, the polymerization rate of this fraction in the three media was 25, 22 and 19 nucleotides added/s to the growing chains. Similar experiments in cultures previously inhibited by 0.2g of chloramphenicol/l showed virtually identical behaviour. This confirmed the work of Midgley & Gray (1971), who, by a different approach, showed that the polymerization rate of rRNA in steadily growing and chloramphenicol-inhibited cultures of E. coli at 37 degrees C was essentially constant at about 22 nucleotides added/s. It was thus confirmed that the rate of polymerization of at least the rRNA fraction in E. coli is virtually unaffected by the nature of the growth medium and therefore by bacterial growth rate.
对添加了0.1g利福平/升后受到抑制的大肠杆菌M.R.E.600培养物中稳定核糖核酸（rRNA + tRNA）和不稳定mRNA组分的标记动力学进行了研究。标记通过添加[2-(14)C]-或[5-(3)H]-尿嘧啶作为细胞核酸的外源前体来进行。通过DNA-RNA杂交研究，追踪了受抑制培养物中mRNA的合成和降解动力学。尽管在利福平存在下标记的相当一部分mRNA降解为不可杂交的产物，但约25%在蛋白质合成最终停止后仍保持稳定。因此，将利福平抑制培养物中mRNA稳定性的研究结果外推到稳定生长速率下的情况似乎是不明智的，在稳定生长速率下，几乎不存在稳定的信使RNA（如果有的话）。受抑制培养物中RNA的标记动力学表明，从添加利福平到放射性不再进入总细胞核糖核酸的时间间隔是聚合一个rRNA分子所需时间的度量。在37℃下，在肉汤、葡萄糖盐或乳酸盐培养基中生长的培养物中，添加抗生素后，外源[2-(14)C]尿嘧啶进入利福平抑制的细胞并在2 - 3分钟内掺入RNA。将这段时间视为聚合一条完整的23S rRNA链所需的时间，在这三种培养基中该组分的聚合速率为每秒向生长链添加25、22和19个核苷酸。在先前用0.2g氯霉素/升抑制的培养物中进行的类似实验显示出几乎相同的行为。这证实了Midgley和Gray（1971）的工作，他们通过不同的方法表明，在37℃下，大肠杆菌稳定生长和氯霉素抑制培养物中rRNA的聚合速率基本恒定，约为每秒添加22个核苷酸。因此证实，大肠杆菌中至少rRNA组分的聚合速率实际上不受生长培养基性质的影响，因此也不受细菌生长速率的影响。
