Role of carbohydrate and protein in the binding of tissue-type plasminogen activator to the human mannose receptor.
The 175-kDa mannose receptor is one of the receptors that mediates the clearance of tissue-type plasminogen activator (t-PA). The affinity of t-PA for the mannose receptor is much higher than the affinity of other high-mannose-type oligosaccharide-containing glycoproteins. In order to find an explanation for this high affinity, we studied the biochemical interaction of various forms of t-PA with the isolated human mannose receptor in several in vitro binding assays. t-PA showed a high affinity (Ki = 0.2 nM) for the mannose receptor and the interaction could be fully inhibited by mannan or polyclonal antibodies against the mannose receptor. The interaction was not affected by non-glycosylated t-PA. The high affinity differed slightly between t-PAs synthesized by various cell types (range Ki 0.2-0.7 nM) and between various glycoforms of t-PA. No statistically significant difference in affinity between t-PA and t-PA complexed to inhibitors was observed. In contrast to intact t-PA, a trypsin digest of t-PA had a low affinity (Ki = 0.5 microM) for the mannose receptor. Both intact and trypsin digests of the high-mannose-type oligosaccharide-containing glycoproteins ribonuclease B and ovalbumin had a low affinity (Ki 0.5-1.5 microM) for the mannose receptor. We conclude that neither protein-protein interactions, nor the complex-type oligosaccharides and the fucose residue on t-PA contribute significantly to the high-affinity binding of t-PA. We suggest that the conformation of the high-mannose-type oligosaccharide on t-PA is influenced by the protein moiety of t-PA in such a way that the oligosaccharide has a high affinity for the mannose receptor.
Noorman
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Gaubius Laboratory, TNO Prevention and Health, Leiden, The Netherlands.
Barrett-Bergshoeff
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D001667:Binding, Competitive; D001710:Biotin / Q000737:chemistry; D005260:Female; D005643:Fucose; D006023:Glycoproteins / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D037181:Lectins, C-Type; D008351:Mannans / Q000378:metabolism; D000090323:Mannose Receptor; D037241:Mannose-Binding Lectins; D008545:Melanoma / Q000378:metabolism; D009844:Oligosaccharides / Q000737:chemistry / Q000378:metabolism; D010047:Ovalbumin / Q000378:metabolism; D010920:Placenta / Q000737:chemistry; D011247:Pregnancy; D011480:Protease Inhibitors / Q000378:metabolism / Q000494:pharmacology; D011956:Receptors, Cell Surface / Q000378:metabolism*; D012260:Ribonucleases / Q000378:metabolism; D010959:Tissue Plasminogen Activator / Q000737:chemistry* / Q000187:drug effects / Q000378:metabolism*; D014357:Trypsin / Q000378:metabolism; D014407:Tumor Cells, Cultured
D006023:Glycoproteins; D037181:Lectins, C-Type; D008351:Mannans; D000090323:Mannose Receptor; D037241:Mannose-Binding Lectins; D009844:Oligosaccharides; D011480:Protease Inhibitors; D011956:Receptors, Cell Surface; D005643:Fucose; D001710:Biotin; D010047:Ovalbumin; D012260:Ribonucleases; C029600:ribonuclease B; D014357:Trypsin; D010959:Tissue Plasminogen Activator