Ohanin, a Novel Protein from King Cobra Venom, Induces Hypolocomotion and Hyperalgesia in Mice

Abstract

We have identified, purified, and determined the complete amino acid sequence of a novel protein, ohanin from Ophiophagus hannah (king cobra) venom. It is a small protein containing 107 amino acid residues with a molecular mass of 11951.47 ؎ 0.67 Da as assessed by electrospray ionization-mass spectrometry. It does not show similarity to any known families of snake venom proteins and hence is the first member of a new family of snake venom proteins. It shows similarity to PRY and SPRY domain proteins. It is nontoxic up to 10 mg/kg when injected intraperitoneally in mice. Ohanin produced statistically significant and dose-dependent hypolocomotion in mice. In a pain threshold assay, it showed dose-dependent hyperalgesic effect. The ability of the protein to elicit a response at greatly reduced doses when injected intracerebroventricularly as compared with intraperitoneal administration in both the locomotion and hot plate experiments strongly suggests that ohanin acts on the central nervous system. Since the natural abundance of the protein in the venom is low (ϳ1 mg/g), a synthetic gene was constructed and expressed. The recombinant protein, which was obtained in the insoluble fraction in Escherichia coli, was purified under denaturing condition and was refolded. Recombinant ohanin is structurally and functionally similar to native protein as determined by circular dichroism and hot plate assay, suggesting that it will be useful in future structure-function relationship studies.

Lachesis muta (Viperidae) cDNAs Reveal Diverging Pit Viper Molecules and Scaffolds Typical of Cobra (Elapidae) Venoms: Implications for Snake Toxin Repertoire Evolution

“…Conversely, the unpublished ohanin precursor available on databases (AAR07992.2) has its starting codon only 20 positions upstream from the mature N terminus ( Figure 5A). The precursor indicates also a C-terminal processing, since the molecules purified from the venom end 63 residues before the stop codon (Pung et al 2005). This processed C terminus is different from other B30.2 proteins ( Figure 5B); it is shorter and predominantly hydrophobic (see plot in Figure 5A), perhaps suggesting a membrane association during the maturation process.…”

“…For the ohanin-like, since the similarity is high and this class of protein was only recently described (Pung et al 2005), being reported also in lizards (Fry et al 2006), we believe that a common origin is very probable. For the diverging 3FTx, the phylogenetic tree also suggests a common and ancestral origin with Colubridae and Elapidae toxins, but in this case, the evidence is not very strong, since the uPAR (domain 2) also was in the group.…”

“…Ohanin-like is also a Viperidae toxin: Ohanin was first isolated from Ophiophagus hannah (king cobra) and shown to cause hypolocomotion and hyperalgesia in mice (Pung et al 2005). Thai cobrin (P82885), another related protein from Naja kaouthia, was deposited in databases without further publication.…”

Structural and Functional Characterization of a Novel Homodimeric Three-finger Neurotoxin from the Venom of Ophiophagus hannah (King Cobra)

“…1). The liquid chromatography/MS profile of O. hannah venom (44) showed the presence of a 7,535.67 Ϯ 0.60 Da protein, similar to the expected molecular mass of the protein being characterized. This protein was purified from the crude venom using a two-step chromatographic approach.…”

The Toxicogenomic Multiverse: Convergent Recruitment of Proteins Into Animal Venoms

“…In reptile venoms, they are an ancestral type shared between lizard and snake venoms (56). The bioactivity of the lizard form remains to be elucidated, but the isoform characterized from Ophiophagus hannah snake venom produces hypolocomotion and hyperalgesia, perhaps by acting directly on the central nervous system (125). SPRY toxins in stonefish venoms produce lethal hemolysis through pore formation in cell membranes while inducing potent endothelium-dependent hypotension and also irreversibly interfering with neuromuscular function (29,88,146).…”