Discovery of a novel vascular endothelial growth factor (VEGF) with no affinity to heparin in Gloydius tsushimaensis venom

• GtVF with strong vascular permeability enhancement was isolated from Gloydius venom.
• The amino acid sequence of GtVF was determined from its cDNA.
• GtVF exhibited no affinity to heparin unlike other snake venom VEGFs.
• A group of snake venom VEGFs with no (or low) affinity to heparin is shown.
• Vascular permeability enhancing activity of GtVF arises through VEGF receptor-2.

Strong vascular permeability enhancing activity was found only in the venom of Gloydius tsushimaensis, in Tsushima island, Japan, when examined together with the venoms of G. blomhoffii snakes in several areas of Japan and of G. ussuriensis in South Korea. The active protein purified by using Superdex 75 and Mono Q columns showed no affinity to heparin, and migrated on SDS-PAGE with molecular weights of 26 and 13 kDa under nonreducing and reducing conditions, respectively, showing that it exists as homodimer. Its N-terminal amino acid sequence was highly homologous to those of snake venom vascular endothelial growth factors (VEGFs). The sequence of this protein, named GtVF, was inferred from the one base-substituted two cDNAs (438 bp) obtained via the 3′ RACE. The phylogenetic analysis suggested the presence of a new type of snake venom VEGFs including GtVF with no affinity to heparin in addition to the known three types of snake venom VEGFs with high affinity to heparin. Since the vascular permeability enhancement by GtVF was inhibited by the antibody against kinase insert domain-containing receptor (KDR), the vascular permeability enhancing activity of GtVF arises through KDR but without heparin binding.