Nucleotide sequence of crotamine isoform precursors from a single South American rattlesnake ( Crotalus durissus terrificus)

Abstract

A cDNA phage library was constructed from venom glands of a single adult specimen of crotamine-plus Crotalus durissus terri®cus (South American rattlesnake) captured in a known region. Fifteen crotamine positive clones were isolated using a PCR-based screening protocol and sequenced. These complete cDNAs clones were grouped for maximal alignment into six distinct nucleotide sequences. The crotamine cDNAs, with 340±360 bases, encompass open reading frame of 198 nucleotides with 50 and 30 untranslated regions of variable size, signal peptide sequence, one crotamine isoform message, and putative poly(A + ) signal. Of these six di erent crotamine cDNA precursors, two predict the identical amino acid sequence previously described by Laure (1975), and the other four a crotamine isoform precursor where the Leucine residue at position 19 is replaced by isoleucine by a single base change. On the other hand, nucleotide variation was observed in the 50 and 30 untranslated regions, with one interesting variant containing an 18 base pair deletion at the 50 untranslated region which results in the usual ATG initiator being replaced by the rarely used GUG start codon. Comparison by Northern blot analysis of poly(A + ) RNA from venom glands of a crotamine-plus specimen to total and poly(A + ) RNA from a crotamine-minus snake