Isolation and characterization of EMS16, a C-lectin type protein from Echis multisquamatus venom, a potent and selective inhibitor of the alpha2beta1 integrin.
We have isolated and characterized EMS16, a potent and selective inhibitor of the alpha2beta1 integrin, from Echis multisquamatus venom. It belongs to the family of C-lectin type of proteins (CLPs), and its amino acid sequence is homologous with other members of this protein family occurring in snake venoms. EMS16 (M(r) approximately 33K) is a heterodimer composed of two distinct subunits linked by S-S bonds. K562 cells transfected with alpha2 integrin selectively adhere to immobilized EMS16, but not to two other snake venom-derived CLPs, echicetin and alboaggregin B. EMS16 inhibits adhesion of alpha2beta1-expressing cells to immobilized collagen I at picomolar concentrations, and the platelet/collagen I interaction in solution at nanomolar concentrations. EMS16 inhibits binding of isolated, recombinant I domain of alpha2 integrin to collagen in an ELISA assay, but not the interaction of isolated I domain of alpha1 integrin with collagen IV. Studies with monoclonal antibodies suggested that EMS16 binds to the alpha2 subunit of the integrin. EMS16 inhibits collagen-induced platelet aggregation, but has no effect on aggregation induced by other agonists such as ADP, thromboxane analogue (U46619), TRAP, or convulxin. EMS16 also inhibits collagen-induced, but not convulxin-induced, platelet cytosolic Ca(2+) mobilization. In addition, EMS16 inhibits HUVEC migration in collagen I gel. In conclusion, we report a new, potent viper venom-derived inhibitor of alpha2beta1 integrin, which does not belong to the disintegrin family.
Sol Sherry Thrombosis Research Center, Department of Pharmacology, and Department of Physiology, Temple University, School of Medicine, 3400 North Broad Street, Philadelphia, Pennsylvania 19140, USA. cmarcink@nimbus.temple.edu
C
C
Marcinkiewicz
R R
RR
Lobb
M M
MM
Marcinkiewicz
J L
JL
Daniel
J B
JB
Smith
C
C
Dangelmaier
P H
PH
Weinreb
D A
DA
Beacham
S
S
Niewiarowski
D000595:Amino Acid Sequence; D020013:Calcium Signaling / Q000187:drug effects; D002448:Cell Adhesion / Q000187:drug effects; D002465:Cell Movement / Q000187:drug effects; D019281:Dimerization; D004220:Disulfides; D004730:Endothelium, Vascular / Q000187:drug effects; D006801:Humans; D016023:Integrins / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D037102:Lectins / Q000145:classification / Q000494:pharmacology*; D037181:Lectins, C-Type; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000037:antagonists & inhibitors / Q000378:metabolism; D010974:Platelet Aggregation / Q000187:drug effects; D011485:Protein Binding; D038981:Receptors, Collagen; D020539:Sequence Analysis, Protein; D014757:Viper Venoms / Q000737:chemistry*
D004220:Disulfides; C412704:EMS16 lectin, Echis multisquamtus; D016023:Integrins; D037102:Lectins; D037181:Lectins, C-Type; D010446:Peptide Fragments; D038981:Receptors, Collagen; D014757:Viper Venoms