Gu Z, Rogers E J, Lovett P S
Department of Biological Sciences, University of Maryland Baltimore County, Catonsville 21228.
J Bacteriol. 1993 Sep;175(17):5309-13. doi: 10.1128/jb.175.17.5309-5313.1993.
The site of ribosome stalling in the leader of cat transcripts is critical to induction of downstream translation. Site-specific stalling requires translation of the first five leader codons and the presence of chloramphenicol, a sequence-independent inhibitor of ribosome elongation. We demonstrate in this report that a synthetic peptide (the 5-mer) corresponding to the N-terminal five codons of the cat-86 leader inhibits peptidyl transferase in vitro. The N-terminal 2-, 3-, and 4-mers and the reverse 5-mer (reverse amino acid sequence of the 5-mer) are virtually without effect on peptidyl transferase. A missense mutation in the cat-86 leader that abolishes induction in vivo corresponds to an amino acid replacement in the 5-mer that completely relieves peptidyl transferase inhibition. In contrast, a missense mutation that does not interfere with in vivo induction corresponds to an amino acid replacement in the 5-mer that does not significantly alter peptidyl transferase inhibition. Our results suggest that peptidyl transferase inhibition by the nascent cat-86 5-mer peptide may be the primary determinant of the site of ribosome stalling in the leader. A model based on this concept can explain the site specificity of ribosome stalling as well as the response of induction to very low levels of the antibiotic inducer.
氯霉素乙酰转移酶（cat）转录本前导序列中核糖体停滞的位点对于下游翻译的诱导至关重要。位点特异性停滞需要翻译前五个前导密码子并存在氯霉素，氯霉素是一种不依赖序列的核糖体延伸抑制剂。我们在本报告中证明，与cat - 86前导序列的N端五个密码子相对应的合成肽（5聚体）在体外抑制肽基转移酶。N端的2聚体、3聚体和4聚体以及反向5聚体（5聚体的反向氨基酸序列）对肽基转移酶几乎没有影响。cat - 86前导序列中一个在体内消除诱导作用的错义突变对应于5聚体中的一个氨基酸替换，该替换完全解除了肽基转移酶的抑制。相反，一个不干扰体内诱导的错义突变对应于5聚体中的一个氨基酸替换，该替换不会显著改变肽基转移酶的抑制。我们的结果表明，新生的cat - 86 5聚体肽对肽基转移酶的抑制可能是前导序列中核糖体停滞位点的主要决定因素。基于这一概念的模型可以解释核糖体停滞的位点特异性以及诱导对极低水平抗生素诱导剂的反应。
