||Analysis on interaction between archaeal ribosomal stalk proteins and translation elongation factors
Honda, Takayoshi本田, 貴嘉
学位の種類: 博士（理学）. 報告番号: 甲第4322号. 学位記番号: 新大院博（理）甲第421号. 学位授与年月日: 平成29年3月23日
The translation elongation cycle is main event in protein synthesis and this cycle requires alternatively binding of two elongation factors. The multiple copies of the flexible ribosomal stalk protein interact directly with the elongation factors and play a crucial role in these processes. Archaeal ribosomes have three stalk homodimers (aP1)_2 that constitute a heptameric complex with the anchor protein aP0. I investigated the factor binding ability of aP1 proteins assembled onto aP0, by gel-retardation assays. The isolated aP0(aP1)_2(aP1)_2(aP1)_2 complex, as well as the form bound to the E. coli 50S core, as a hybrid 50S particle, interacted strongly with elongation factor aEF2, but weakly with aEF1A. These interactions were disrupted by a point mutation, F107S, at the C-terminus of aP1. To examine the ability of each copy of aP0-associated aP1 to bind to elongation factors, I constructed aP0•aP1 variant trimers, composed of an aP0 mutant and a single (aP1)_2 dimer. Biochemical and quantitative analyses revealed that the resultant three trimers, aP0(aP1)_2＾I, aP0(aP1)_2＾II, and aP0(aP1)_2＾III, individually bound two molecules of aEF2, suggesting that each copy of the aP1 C-terminal region in the aP0•aP1 trimers can bind tightly to aEF2. Interestingly, the unstable binding of aEF1A to each of the three aP0•aP1 trimers was remarkably stabilized in the presence of aEF2. This stabilization of the aEF1A binding to the stalk complex may contribute to promotion of an efficient translation elongation cycle.