Cloning, Tissue-specific Expression And Evolutional Analysis Of CNBP And EIF-5A In Grass Carp

Ctenopharyngodon idella cellular nucleic acid-binding protein(CNBP) and eukaryotic translation initiation factor 5A(eIF-5A) had been cloned from the liver and kidney grass carp cDNA library.The full-lenghth grass carp CNBP was 1948bp,which contained a 5' untranslated region of 33bp and a 3' untranslated region of 1423bp.The open reading frame was 492bp,which could code a 163 amino acid polypeptide. Grass carp CNBP contained seven CCHC zinc finger domains,nuclear located signal and an RGG box,which was highly conserved in fish.Compared with human and other vertebrates,25 amino acid residues was subsituted in grass carp CNBP,the fifth glycine residue at the third zinc finger domain was substituted with histidine and 6-14 amino acid residues were absent between the first and the second zinc finger in the grass carp CNBP.Nevertheless,the grass carp CNBP was not in the positive Darwin selection(ω≤1) by adaptive evolution analysis.Therefore,new function was not generated by structural difference of CNBP.These indicated that CNBP was in the neutral selection.RT-PCR experiment indicted that in the normal liver,intestine, kidney,spleen,heart,muscle,gill and brain of grass carp,the CNBP had the highest expressional level in heart,the brain and gill ranked secondly,and weakly in kidney.The full length cDNA sequence of grass carp eIF-5A was 2249bp,which contained a 5' untranslated region of 41bp and a 3' untranslated region of 1743bp. The open reading frame was 465bp,which could code a 155 amino acid polypeptide with a molecular mass of 16.856 ku and pI 5.18.Grass carp eIF-5A contains the unusual basic amino acid hypusine that conserved in eIF-5A family at the N-terminal region and a leucine-rich stretch at the C-terminal region.And its complete amino acid sequence shared highly identity with eIF-5A from human and other species,only several amino acid residues were substituted.Adaptive evolution analysis also indicated that eIF-5A was not only conserved in structure but also in function.eIF-5A is constantly expressed among the heart,brain,muscle,gill,spleen,liver,intestine and kidney of grass carp,as well as in infected tissues by Poly I:C.