LEISHMANIA-BRAZILIENSIS, MOLECULAR CHARACTERIZATION OF AN ELONGATION-FACTOR 1-ALPHA GENE

The elongation factor EF-1 alpha is one of the most studied components of the translation machinery owing to its abundance and possible role in other cellular functions. EF-1 alpha mediates the correct coupling of the aminoacyl-tRNA on the A site of the ribosome in a GTP-dependen t reaction. We have previously described an EF-1 alpha DNA sequence in Leishmania amazonensis, pLEF11 (accession No. M92653), using PCR. In this paper we describe the DNA sequence and genomic organization of L. braziliensis EF-1 alpha gene. Southern blot analysis revealed that EF -1 alpha is organized as a 2 kb tandem repeat. The pLEF11 probe recogn ized a 1.8 kb mRNA from promastigotes in Northern blots. A clone conta ining the first copy and a half of the EF-1 alpha tandem repeat was is olated by screening a L. braziliensis genomic library. Southern blot a nalysis showed that the isolated clone (lambda 2.2) presented the same hybridization profile as that of a genomic blot. The partial sequenci ng of clone lambda 2.2 spans 2959 nucleotides in length, which has two open reading frames separated by a putative non-coding region. The nu cleotide and the predicted peptide sequence of the first coding region presented approximately 80% identity with other eukaryotic EF-1 alpha genes. The sequence also displayed the four consensus motifs correspo nding to the GTP-binding site (G1, G2, G3 and G4). Computer analysis o f the sequence of both coding regions revealed three divergent nucleot ides, which generated two changes at the amino acid level. One was fou nd to be located in the G2 domain. The non-coding region of the EF-1 a lpha gene sequence showed potential regulatory elements such as polypy rimidine tracks, chi-homologous sequences and stem-loop forming sequen ces. (C) 1997 Elsevier Science B.V.