The shape of thrombomodulin and interactions with thrombin as determined by electron microscopy.
Studies have been carried out to investigate aspects of the structure of thrombomodulin, an endothelial cell glycoprotein that binds thrombin and accelerates both the thrombin-dependent activation of protein C and the inhibition of antithrombin III. We have determined the shape of SolulinTM, a soluble recombinant form of human thrombomodulin missing the transmembrane and cytoplasmic domains, by electron microscopy of preparations rotary-shadowed with tungsten. Solulin appears to be an elongated molecule about 20 nm long that has a large nodule at one end and a smaller nodule near the other end from which extends a thin strand. About half of the molecules form bipolar dimers apparently via interactions between these thin strands. Electron microscopy of complexes formed between Solulin and human alpha-thrombin revealed that a single thrombin molecule appears to bind to the smaller nodule of Solulin, suggesting that this region contains the epidermal growth factor-like domains 5 and 6. Epidermal growth factor-like domains 1-4 comprise the connector between the small and large nodule, which is the lectin-like domain; the thin strand at the other end of the molecule is the carbohydrate-rich region. With chondroitin sulfate-containing soluble thrombomodulin produced from either human melanoma cells Bowes or Chinese hamster ovary cells, a higher percentage of molecules bound thrombin and, in some cases, two thrombin molecules were attached to one soluble thrombomodulin in approximately the same region. These structural studies provide insight into the structure of thrombomodulin and its interactions with thrombin as well as aspects of the mechanisms of its actions.
Weisel
J W
JW
Department of Cell and Developmental Biology, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104-6058, USA.
Nagaswami
C
C
Young
T A
TA
Light
D R
DR
D000818:Animals; D016466:CHO Cells; D002809:Chondroitin Sulfates / Q000378:metabolism; D006224:Cricetinae; D018360:Crystallography, X-Ray; D006801:Humans; D008854:Microscopy, Electron; D008956:Models, Chemical; D012995:Solubility; D013917:Thrombin / Q000378:metabolism*; D018180:Thrombomodulin / Q000378:metabolism / Q000648:ultrastructure*
D018180:Thrombomodulin; D002809:Chondroitin Sulfates; D013917:Thrombin