Isolation of the pituitary gonadotrophic alpha-subunit hormone of the giant amazonian fish: pirarucu (Arapaima gigas)

摘要

The cDNAs of the alpha-subunit of the pituitary gonadotrophic hormones (GTH alpha) of fish of the order Osteoglossiformes or the superorder Osteoglossomorpha have never been sequenced. For a better understanding the phylogenetic diversity and evolution of PGH alpha in fish and for future biotechnological synthesis of the gonadotrophic hormones (ag-FSH and ag-LH), of Arapaima gigas, one of the largest freshwater fishes of the world, its GTH alpha cDNA was synthesized by reverse transcriptase and the polymerase chain reaction starting from total pituitary RNA. The ag-GTH alpha-subunit was found to be encoded by 348 bp, corresponding to a protein of 115 amino acids, with a putative signal peptide of 24 amino acids and a mature peptide of 91 amino acids. Ten cysteine residues, responsible for forming 5 disulfide linkages, 2 putative N-linked glycosylation sites and 3 proline residues, were found to be conserved on the basis of the known sequences of vertebrate gonadotrophic hormones. Phylogenetic analysis, based on the amino acid sequences of 38 GTH alpha-subunits, revealed the highest identity of A. gigas with members of the Acipenseriformes, Anguilliformes, Siluriformes and Cypriniformes (87.1-89.5 %) and the lowest with Gadiformes and Cyprinodontiformes (55.0 %). The obtained phylogenetic tree agrees with previous analysis of teleostei, since A. gigas, of the order of Osteoglossiformes, appears as the sister group of Clupeocephala, while Elopomorpha forms the most basal group of all other teleosts.