Molecular Cloning and Characterization of theCaenorhabditis elegansElongation Factor 2 Gene (eft-2)

摘要

ABSTRACTACaenorhabditis elegansλZAP cDNA library was screened using a fragment amplified from highly conserved regions of the mammalian andDrosophilaelongation factor 2 (EF-2). Two types of cDNA clones were obtained, corresponding to two mRNA species with 3′-untranslated regions of 60 and 115 nucleotides, both encoding identical polypeptides. Sequence analysis of these clones and comparisons with hamster andDrosophilaEF-2 sequences suggests that they encodeC. elegansEF-2. Clone pCef6A, encoding the entireC. elegansEF-2 mRNA sequence including 45 nucleotides of 5′-untranslated region, contains a 2,556-bp open reading frame which predicts a polypeptide of 852 amino acid residues (Mr94,564). The deduced amino acid sequence is greater than 80 identical to that of mammalian andDrosophilaEF-2. Conserved sequence segments shared among a variety of GTP-binding proteins are found in the amino-terminal region. The carboxy-terminal half contains segments unique to EF-2 and its prokaryotic homolog, EF-G, as well as the histidyl residue which is ADP-ribosylated by diphtheria toxin. TheC. elegansprotein contains a 12-amino-acid insertion between positions 90 and 100, and a 13-amino-acid deletion between positions 237 and 260, relative to hamster EF-2. Partial sequencing of a genomic clone encoding the entireC. elegansEF-2 gene (namedeft-2) has so far revealed two introns of 48 and 44 bp following codons Gln-191 and Gln-250, respectively. Southern and Northern blot analyses indicate thateft-2is a single-copy gene and encodes a 3-kb mRNA species which is present throughout nematode develop