title: Matrix localization of tissue factor pathway inhibitor-2/matrix-associated serine protease inhibitor (TFPI-2/MSPI) involves arginine-mediated ionic interactions with heparin and dermatan sulfate: heparin accelerates the activity of TFPI-2/MSPI toward plasmin.
abstract: Human tissue factor pathway inhibitor-2 (TFPI-2)/matrix-associated serine protease inhibitor (MSPI), a Kunitz-type serine protease inhibitor, inhibits plasmin, trypsin, chymotrypsin, plasma kallikrein, cathepsin G, and factor VIIa-tissue factor complex. The mature protein has a molecular mass of 32-33 kDa, but exists in vivo as two smaller, underglycosylated species of 31 and 27 kDa. TFPI-2/MSPI triplet is synthesized and secreted by a variety of cell types that include epithelial, endothelial, and mesenchymal cells. Because the majority (75-90%) of TFPI-2/MSPI is associated with the extracellular matrix (ECM), we examined which components of the ECM bind TFPI-2/MSPI. We found that TFPI-2/MSPI bound specifically to heparin and dermatan sulfate. Interaction of these two glycosaminoglycans (GAGs) with TFPI-2/MSPI involved one or more common protein domains, as evidenced by cross-competition experiments. However, binding affinity for TFPI-2/MSPI with heparin was 250-300 times greater than that for TFPI-2/MSPI with dermatan sulfate. Binding of TFPI-2/MSPI to GAGs was inhibited by NaCl or arginine but not by glucose, mannose, galactose, 6-aminohexanoic acid, or urea, suggesting that arginine-mediated ionic interactions participate in the GAG binding of TFPI-2/MSPI. This supposition was supported by the observation that only NaCl or arginine could elute the TFPI-2/MSPI protein triplet from an ECM derived from human dermal fibroblasts. Reduced TFPI-2/MSPI did not bind to heparin, suggesting that proper disulfide pairings and conformation are essential for matrix binding. To determine whether heparin modulates the activity of TFPI-2/MSPI, we determined the rate of inhibition of plasmin by the inhibitor with and without heparin and found that TFPI-2/MSPI is more active in the presence of heparin. Collectively, our results demonstrate that conformation-dependent arginine-mediated ionic interactions are responsible for the TFPI-2/MSPI triplet binding to fibroblast ECM, heparin, and dermatan sulfate and that heparin augmented the rate of inhibition of plasmin by TFPI-2/MSPI.
authors.affiliation: Department of Neurosurgery, The University of Texas M.D. Anderson Cancer Center, Houston, Texas, 77030, USA.
authors.forename: Y
authors.initials: Y
authors.lastname: Liu
authors.forename: S M
authors.initials: SM
authors.lastname: Stack
authors.forename: S S
authors.initials: SS
authors.lastname: Lakka
authors.forename: A J
authors.initials: AJ
authors.lastname: Khan
authors.forename: D T
authors.initials: DT
authors.lastname: Woodley
authors.forename: J S
authors.initials: JS
authors.lastname: Rao
authors.forename: C N
authors.initials: CN
authors.lastname: Rao
mesh_terms: D001120:Arginine / Q000378:metabolism*; D001667:Binding, Competitive; D056649:Cathepsin G; D002403:Cathepsins / Q000037:antagonists & inhibitors; D002918:Chymotrypsin / Q000378:metabolism; D003546:Cysteine Endopeptidases / Q000378:metabolism; D003871:Dermatan Sulfate / Q000378:metabolism* / Q000494:pharmacology; D005341:Fibrinolysin / Q000037:antagonists & inhibitors*; D006023:Glycoproteins / Q000378:metabolism / Q000494:pharmacology*; D006493:Heparin / Q000378:metabolism* / Q000494:pharmacology; D006801:Humans; D007610:Kallikreins / Q000037:antagonists & inhibitors; D007700:Kinetics; D009363:Neoplasm Proteins; D011257:Pregnancy Proteins / Q000378:metabolism / Q000494:pharmacology*; D012697:Serine Endopeptidases; D015842:Serine Proteinase Inhibitors / Q000494:pharmacology*; D014357:Trypsin / Q000378:metabolism
chemical_list: D006023:Glycoproteins; D009363:Neoplasm Proteins; D011257:Pregnancy Proteins; D015842:Serine Proteinase Inhibitors; C021240:tissue-factor-pathway inhibitor 2; D003871:Dermatan Sulfate; D006493:Heparin; D001120:Arginine; D002403:Cathepsins; D007610:Kallikreins; D012697:Serine Endopeptidases; D002918:Chymotrypsin; C534094:CTSG protein, human; D056649:Cathepsin G; D014357:Trypsin; D005341:Fibrinolysin; D003546:Cysteine Endopeptidases; C030910:cancer procoagulant