CROTAMINE AND CROTASIN, TWO EVOLUTIONARY RELATED PEPTIDES FROM THE Crotalus durissus terrificus (SOUTH AMERICAN RATTLESNAKE)

RÁDIS-BAPTISTA, G.(1), HAYASHÍ, M.A.F.(2), GREGO, K.F.(3), OLIVEIRA, E.B.(4), YAMANE, T.(1)

(1)Molecular Toxinology Laboratory, (2)Biochemistry and Biophysics Laboratory, (3)Herpetology Laboratory, Institute Butantan, São Paulo, SP, (4)Biochemistry Department, Faculty of Medicine, USP, RibeirãoPreto, SP.

Crotamine (myotoxin) is a small basic peptide (MW 4400, pl > 9.5), widely distributed among venoms of the genera Crotalus from the prairies of the Western United States to the pampas of South America. However, its presence/absence varies according to the subspecies and the geographic locality of a given species (Schenberg, S., 1959. Science 129:1361, Ownby, C.L., 1998. J. Toxicol. 17:213). In order to address the evolutionary relationship between crotamine-positive and -negative variants, their genomic structures were studied, to verify the structural organization of crotamine gene and why it is not transcribed in the venom glands of crotamine-negative specimens. Genomic DNA, Crt-p1, with the size of 1.8 kb, encoding the precursor of crotamine was isolated from the liver of crotamine-positive specimen, but we were unable to find similar gene in the crotamine-negative one. Instead, a 2.5 kb size gene, designated crotasin, was isolated, paralogous to crotamine gene, presenting virtually identical overall structural organization: three exons separated by two introns, a long phase-1 and a short phase-2 introns. Crotasin gene, Cts-p2, which happens to occur in both crotamine-positive and -negative ones, when compared to Crt-p1 gene, shows a considerable mutations involving a high nucleotide substitutions (134 bases), as well as deletions (23 bases) and an insertion of 892 bases, due partly to the duplication (760 bases) of intron segments. Base substitutions occur mostly in the crotamine-coding region, leading to a hypermutated peptide (only 11 out of 42 amino acids are preserved), in contrast to a highly conserved signal peptide sequence and equally conserved upstream and downstream gene flanking regions. One striking fact is that all 6 cysteine residues are preserved maintaining, probably, the same fold. Crotasin is expressed in various tissues, and at a high concentration in pancreas. The probable function of this novel peptide is under investigation.

Supported by: FAPESP

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GANDHI RÁDIS BAPTISTA, Avenida Miruna, 1808, São Paulo, SP, CEP: 04084-006, Brasil, Email: radisbra@yahoo.com