Initiation factor eIF5B catalyzes second GTP-dependent step in eukaryotic translation initiation

^*Laboratory of Gene Regulation and Development, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892-2716; ^‡Department of Microbiology and Immunology, State University of New York Health Science Center, Brooklyn, NY 11203; and ^§A. N. Belozersky Institute of Physico-Chemical Biology, Moscow State University, Moscow, Russia

Edited by Harry F. Noller, University of California, Santa Cruz, CA, and approved October 31, 2002 (received for review September 19, 2002)

Abstract

Initiation factors IF2 in bacteria and eIF2 in eukaryotes are GTPases that bind Met-tRNA $\text{[math]}$ to the small ribosomal subunit. eIF5B, the eukaryotic ortholog of IF2, is a GTPase that promotes ribosomal subunit joining. Here we show that eIF5B GTPase activity is required for protein synthesis. Mutation of the conserved Asp-759 in human eIF5B GTP-binding domain to Asn converts eIF5B to an XTPase and introduces an XTP requirement for subunit joining and translation initiation. Thus, in contrast to bacteria where the single GTPase IF2 is sufficient to catalyze translation initiation, eukaryotic cells require hydrolysis of GTP by both eIF2 and eIF5B to complete translation initiation.

Footnotes

↵ † J.H.L. and T.V.P. contributed equally to this work.
↵ ¶ To whom correspondence should be addressed. E-mail: tdever{at}box-t.nih.gov.
This paper was submitted directly (Track II) to the PNAS office.
Abbreviations:
1. IF, initiation factor
2. eIF, eukaryotic IF
3. EF, elongation factor
4. MP, methionyl-puromycin