Nowotny V, Nierhaus K H
Max-Planck-Institut für Molekulare Genetik, Abteilung Wittmann, Berlin-Dahlem, West Germany.
Biochemistry. 1988 Sep 6;27(18):7051-5. doi: 10.1021/bi00418a057.
A protein which initiates assembly of ribosomes is defined as a protein which binds to the respective rRNA without cooperativity (i.e., without the help of other proteins) during the onset of assembly and is essential for the formation of active ribosomal subunits. The number of proteins binding without cooperativity was determined by monitoring the reconstitution output of active particles at various inputs of 16S rRNA, in the presence of constant amounts of 30S-derived proteins (TP30): This showed that only two of the proteins of the 30S subunit are assembly-initiator proteins. These two proteins are still present on a LiCl core particle comprising 16S rRNA and 12 proteins (including minor proteins). The 12 proteins were isolated, and a series of reconstitution experiments at various levels of rRNA excess demonstrated that S4 and S7 are the initiator proteins. Pulse-chase experiments performed during the early assembly with 14C- and 3H-labeled TP30 and the determination of the 14C/3H ratio of the individual proteins within the assembled particles revealed a bilobal structure of the 30S assembly: A group of six proteins headed by S4 (namely, S4, S20, S16, S15, S6, and S18) resisted the chasing most efficiently (S4 assembly domain). None of the proteins depending on S7 during assembly were found in this group but rather in a second group with intermediate chasing stability [S7 assembly domain; consisting of S7, S9, (S8), S19, and S3]. A number of proteins could be fully chased during the early assembly and therefore represent "late assembly proteins" (S10, S5, S13, S2, S21, S1). These findings fit well with the 30S assembly map.(ABSTRACT TRUNCATED AT 250 WORDS)
一种启动核糖体组装的蛋白质被定义为在组装开始时能与相应的核糖体RNA（rRNA）非协同性结合（即无需其他蛋白质的帮助）且对活性核糖体亚基的形成至关重要的蛋白质。通过在16S rRNA的不同输入量以及恒定数量的30S衍生蛋白质（TP30）存在的情况下监测活性颗粒的重组输出，来确定非协同性结合的蛋白质数量：这表明30S亚基中只有两种蛋白质是组装起始蛋白。这两种蛋白质仍存在于包含16S rRNA和12种蛋白质（包括次要蛋白质）的氯化锂核心颗粒上。分离出这12种蛋白质，并在不同程度的rRNA过量情况下进行了一系列重组实验，结果表明S4和S7是起始蛋白。在组装早期用14C和3H标记的TP30进行脉冲追踪实验，并测定组装颗粒中各个蛋白质的14C/3H比率，揭示了30S组装的双叶结构：以S4为首的一组六种蛋白质（即S4、S20、S16、S15、S6和S18）最有效地抵抗追踪（S4组装结构域）。在组装过程中依赖S7的蛋白质均不在该组中，而是在具有中等追踪稳定性的第二组中（S7组装结构域；由S7、S9、（S8）、S19和S3组成）。许多蛋白质在组装早期可以被完全追踪，因此代表“后期组装蛋白质”（S10、S5、S13、S2、S21、S1）。这些发现与30S组装图谱非常吻合。（摘要截短于250字）
