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首页> 外文期刊>The Journal of biological chemistry >Plant Cap-binding Complexes Eukaryotic Initiation Factors eIF4F and eIFISO4F
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Plant Cap-binding Complexes Eukaryotic Initiation Factors eIF4F and eIFISO4F

机译：植物帽结合复合物真核起始因子EIF4F和EIFISO4F

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The initiation of translation in eukaryotes requires a suite of eIFs that include the cap-binding complex, eIF4F. eIF4F is comprised of the subunits eIF4G and eIF4E and often the helicase, eIF4A. The eIF4G subunit serves as an assembly point for other initiation factors, whereas eIF4E binds to the 7-methyl guanosine cap of mRNA. Plants have an isozyme form of eIF4F (eIFiso4F) with comparable subunits, eIFiso4E and eIFiso4G. Plant eIF4A is very loosely associated with the plant cap-binding complexes. The specificity of interaction of the individual subunits of the two complexes was previously unknown. To address this issue, mixed complexes (eIF4E-eIFiso4G or eIFiso4E-eIF4G) were expressed and purified from Escherichia coli for biochemical analysis. The activity of the mixed complexes in in vitro translation assays correlated with the large subunit of the respective correct complex. These results suggest that the eIF4G or eIFiso4G subunits influence translational efficiency more than the cap-binding subunits. The translation assays also showed varying responses of the mRNA templates to eIF4F or eIFiso4F, suggesting that some level of mRNA discrimination is possible. The dissociation constants for the correct complexes have KD values in the subnanomolar range, whereas the mixed complexes were found to have KD values in the ～10 nm range. Displacement assays showed that the correct binding partner readily displaces the incorrect binding partner in a manner consistent with the difference in KD values. These results show molecular specificity for the formation of plant eIF4F and eIFiso4F complexes and suggest a role in mRNA discrimination during initiation of translation.
机译：真核生物中翻译的发起需要一套包括帽结合复合物的EIF4F套件。 EIF4F由亚基EIF4G和EIF4E组成，通常是螺旋酶，EIF4A。 EIF4G亚基用作其他启动因子的组装点，而EIF4E与mRNA的7-甲基胍帽结合。植物具有同工酶形式的EIF4F（EIFISO4F），具有可比较的亚基，EIFISO 4E和EIFISO4G。植物EIF4A与植物帽结合复合物非常松散地相关。两种复合物的个体亚基相互作用的特异性先前未知。为了解决这个问题，从大肠杆菌表达并纯化混合复合物（EIF4E-EIFISO4G或EIFISO4E-EIF4G以进行生化分析。在体外转化测定中的混合络合物的活性与各种正确复合物的大亚基相关。这些结果表明，EIF4G或EIFISO4G亚基比帽绑定亚基更多地影响平移效率。翻译测定还显示出MRNA模板对EIF4F或EIFISO4F的改变响应，表明可以实现某些水平的mRNA歧视。用于正确复合物的解离常数在亚甲醛系列中具有KD值，而发现混合络合物在约10nm范围内具有Kd值。位移测定表明，正确的结合伴侣以与KD值的差异一致的方式易于取代不正确的结合伴侣。这些结果表明，植物EIF4F和EIFISO4F复合物形成的分子特异性，并表明在翻译启动期间mRNA歧视中的作用。

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