Spencer Angela C, Spremulli Linda L
Department of Chemistry, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599-3290, USA.
Biochim Biophys Acta. 2005 Jun 15;1750(1):69-81. doi: 10.1016/j.bbapap.2005.03.009. Epub 2005 Apr 7.
Bovine mitochondrial translational initiation factor 2 (IF-2(mt)) is organized into four domains, an N-terminal domain, a central G-domain and two C-terminal domains. These domains correspond to domains III-VI in the six-domain model of Escherichia coli IF-2. Variants in IF-2(mt) were prepared and tested for their abilities to bind the small (28S) subunit of the mitochondrial ribosome. The binding of wild-type IF-2(mt) was strong (K(d) approximately 10-20 nM) and was not affected by fMet-tRNA. Deletion of the N-terminal domain substantially reduced the binding of IF-2(mt) to 28S subunits. However, the addition of fMet-tRNA stimulated the binding of this variant at least 2-fold demonstrating that contacts between fMet-tRNA and IF-2(mt) can stabilize the binding of this factor to 28S subunits. No binding was observed for IF-2(mt) variants lacking the G-domain which probably plays a critical role in organizing the structure of IF-2(mt). IF-2(mt) contains a 37-amino acid insertion region between domains V and VI that is not found in the prokaryotic factors. Mutations in this region caused a significant reduction in the ability of the factor to promote initiation complex formation and to bind 28S subunits.
牛线粒体翻译起始因子2（IF-2(mt)）由四个结构域组成，一个N端结构域、一个中央G结构域和两个C端结构域。这些结构域对应于大肠杆菌IF-2六结构域模型中的结构域III-VI。制备了IF-2(mt)的变体，并测试它们结合线粒体核糖体小（28S）亚基的能力。野生型IF-2(mt)的结合力很强（解离常数K(d)约为10-20 nM），且不受甲硫氨酰-tRNA的影响。N端结构域的缺失显著降低了IF-2(mt)与28S亚基的结合。然而，添加甲硫氨酰-tRNA可使该变体的结合至少增强2倍，这表明甲硫氨酰-tRNA与IF-2(mt)之间的相互作用可稳定该因子与28S亚基的结合。对于缺乏G结构域的IF-2(mt)变体未观察到结合，G结构域可能在IF-2(mt)的结构组织中起关键作用。IF-2(mt)在结构域V和VI之间含有一个37个氨基酸的插入区域，这在原核因子中未发现。该区域的突变导致该因子促进起始复合物形成及结合28S亚基的能力显著降低。
