cDNA sequence analysis and expression of cardiotoxin V and a new cardiotoxin VII from Naja naja atra (Taiwan cobra)

The cDNAs encoding cardiotoxin V and a new cardiotoxin VII were constructed from the cellular RNA isolated from the venom glands of Naja naja atra by reverse transcription-polymerase chain reaction. Although 95% nucleotide sequence homology was observed with the two cardiotoxins, there were nine amino-acid substitutions between cardiotoxin V and cardiotoxin VII. The cardiotoxins were subcloned into the expression vector pET 20b(+) and transformed into BL21(DE3) E. coli strain. The expressed protein was isolated from the inclusion bodies of E. coli, and purified by reverse-phase high-performance liquid chromatography. The purified recombinant cardiotoxin showed immunoreactivity with anti-cardiotoxin III antibodies as revealed by immunoblot analysis.