We report the isolation and amino acid sequences of six novel dimeric disintegrins from the venoms of Vipera lebetina obtusa (VLO), V. berus (VB), V. ammodytes (VA), Echis ocellatus (EO) and Echis multisquamatus (EMS). Disintegrins VLO4, VB7, VA6 and EO4 displayed the RGD motif and inhibited the adhesion of K562 cells, expressing the integrin α5β1 to immobilized fibronectin. A second group of dimeric disintegrins (VLO5 and EO5) had MLD and VGD motifs in their subunits and blocked the adhesion of the α4β1 integrin to vascular cell adhesion molecule 1 with high selectivity. On the other hand, disintegrin EMS11 inhibited both α5β1 and α4β1 integrins with almost the same degree of specificity. Comparison of the amino acid sequences of the dimeric disintegrins with those of other disintegrins by multiple-sequence alignment and phylogenetic analysis, in conjunction with current biochemical and genetic data, supports the view that the different disintegrin subfamilies evolved from a common ADAM (a disintegrin and metalloproteinase-like) scaffold and that structural diversification occurred through disulphide bond engineering.
- disulphide bond
- protein sequence
- snake venom protein
Abbreviations used: ADAM, a disintegrin and metalloproteinase-like; CHO, Chinese-hamster ovary; EMS, disintegrin from the venom of Echis multisquamatus; EO, VA, VB, VLO, disintegrins from the venoms of Echis ocellatus, Vipera ammodytes, V. berus and V. lebetina obtusa respectively; HBSS, Hanks balanced salt solution; MALDI–TOF-MS, matrix-assisted laser-desorption ionization–time-of-flight mass spectrometry; PE, pyridylethylated; TFA, trifluoroacetic acid; VCAM, vascular cell adhesion molecule.
- The Biochemical Society, London ©2003