Mutational analysis of the mammalian translation initiation factor eIF-4A

Abstract

eIF-4A is a eukaryotic translation initiation factor and DEAD box RNA helicase that is thought to be responsible for the melting of secondary structure in the 5$sp prime$ untranslated region of messenger RNAs to facilitate ribosome binding. A mutational analysis of eIF-4A revealed that the ATPase A motif (AXXXXGKT) is involved in ATP binding, the ATPase B motif (DEAD) is implicated in ATP hydrolysis, the SAT region is essential for RNA unwinding, and the HRIGRXXR region is required for ATP hydrolysis-dependent RNA binding. Furthermore, defective eIF-4A mutants exhibit a strong dominant negative effect on in vitro translation of several mRNAs, including those translated by a cap-independent internal initiation mechanism. It is demonstrated that eIF-4A functions primarily as a subunit of eIF-4F, and singular eIF-4A is required to recycle through the eIF-4F during translation. Accordingly, eIF-4F appears to be required for cap-dependent and internal initiation of translation