Department of Molecular Structural Biology, Institute of Microbiology and Genetics, Georg-August University Göttingen, Göttingen, Germany.
PLoS One. 2010 Sep 16;5(9):e12784. doi: 10.1371/journal.pone.0012784.
The multi-subunit eukaryotic initiation factor3 (eIF3) plays a central role in the initiation step of protein synthesis in eukaryotes. One of its large subunits, eIF3b, serves as a scaffold within eIF3 as it interacts with several other subunits. It harbors an RNA Recognition Motif (RRM), which is shown to be a non-canonical RRM in human as it is not capable to interact with oligonucleotides, but rather interacts with eIF3j, a sub-stoichiometric subunit of eIF3.
We have analyzed the high-resolution crystal structure of the eIF3b RRM domain from yeast. It exhibits the same fold as its human ortholog, with similar charge distribution on the surface interacting with the eIF3j in human. Thermodynamic analysis of the interaction between yeast eIF3b-RRM and eIF3j revealed the same range of enthalpy change and dissociation constant as for the human proteins, providing another line of evidence for the same mode of interaction between eIF3b and eIF3j in both organisms. However, analysis of the surface charge distribution of the putative RNA-binding β-sheet suggested that in contrast to its human ortholog, it potentially could bind oligonucleotides. Three-dimensional positioning of the so called "RNP1" motif in this domain is similar to other canonical RRMs, suggesting that this domain might indeed be a canonical RRM, conferring oligonucleotide binding capability to eIF3 in yeast. Interaction studies with yeast total RNA extract confirmed the proposed RNA binding activity of yeast eIF3b-RRM.
We showed that yeast eIF3b-RRM interacts with eIF3j in a manner similar to its human ortholog. However, it shows similarities in the oligonucleotide binding surface to canonical RRMs and interacts with yeast total RNA. The proposed RNA binding activity of eIF3b-RRM may help eIF3 to either bind to the ribosome or recruit the mRNA to the 43S pre-initiation complex.
多亚基真核起始因子 3（eIF3）在真核生物蛋白质合成的起始步骤中发挥核心作用。其一个大亚基 eIF3b 在 eIF3 内作为支架，与其他几个亚基相互作用。它具有 RNA 识别基序（RRM），在人类中被证明是一种非典型的 RRM，因为它不能与寡核苷酸相互作用，而是与 eIF3j 相互作用，eIF3j 是 eIF3 的亚化学计量亚基。
我们分析了来自酵母的 eIF3b RRM 结构域的高分辨率晶体结构。它表现出与人类同源物相同的折叠，表面的电荷分布相似，与人类中的 eIF3j 相互作用。酵母 eIF3b-RRM 与 eIF3j 之间相互作用的热力学分析显示出相同的焓变和离解常数范围，为这两种生物中 eIF3b 和 eIF3j 之间相同的相互作用模式提供了另一条证据。然而，对假定的 RNA 结合β-折叠的表面电荷分布的分析表明，与人类同源物相比，它可能潜在地能够结合寡核苷酸。该结构域中所谓的“RNP1”基序的三维定位与其他典型的 RRMs 相似，表明该结构域实际上可能是一个典型的 RRM，赋予酵母 eIF3 寡核苷酸结合能力。与酵母总 RNA 提取物的相互作用研究证实了酵母 eIF3b-RRM 提出的 RNA 结合活性。
我们表明，酵母 eIF3b-RRM 以与人类同源物相似的方式与 eIF3j 相互作用。然而，它在与典型 RRMs 结合寡核苷酸的表面上显示出相似性，并与酵母总 RNA 相互作用。eIF3b-RRM 的提出的 RNA 结合活性可能有助于 eIF3 结合核糖体或招募 mRNA 到 43S 起始前复合物。
