The Kringle V-Protease Domain Is a Fibrinogen Binding Region within Apo(a)

Song Xue; Edwin L. Madison; Lindsey A. Miles

doi:10.1055/s-0037-1616056

Thrombosis and Haemostasis, Table of Contents

Thromb Haemost 2001; 86(05): 1229-1237
DOI: 10.1055/s-0037-1616056

Review Article

Schattauer GmbH

The Kringle V-Protease Domain Is a Fibrinogen Binding Region within Apo(a)

Authors

Song Xue

¹The Scripps Research Institute, Department of Vascular Biology, La Jolla, California, USA
Edwin L. Madison^*

¹The Scripps Research Institute, Department of Vascular Biology, La Jolla, California, USA
Lindsey A. Miles

¹The Scripps Research Institute, Department of Vascular Biology, La Jolla, California, USA

Abstract

Summary

Lp(a) binds directly to fibrin and competes for the interaction of plasminogen with this substrate. This competition may play a role in the proatherothrombogenic consequences of high Lp(a) levels. Previous studies by us and others showed that apo(a) Kringle IV-10 competes for the interaction of Lp(a) with plasmin-treated fibrinogen. However, kringle IV-10 cannot account for the entire high affinity interaction of Lp(a) with fibrinogen. Therefore, we tested the hypothesis that the apo(a) kringle V protease-like domain (KV-PD) could interact with plasmin-treated fibrinogen. We cloned the apo(a) KV-PD region from a human liver cDNA library. Fusion apo(a) KV-PD was expressed in COS 7 cells and purified from the conditioned media. Western blotting of the apo(a) KV-PD protein revealed two bands migrating with apparent molecular weights of 45K and 48K. When fusion apo(a) KV-PD was treated with O-glycosidase and neuraminidase, the higher molecular weight band disappeared suggesting that the apo(a) KV-PD was O-glycosylated. Apo(a) KV-PD bound to plasmin-treated fibrinogen in a dose-dependent fashion. An EC₅₀ of 3.9 ± 0.2 μM was determined for this interaction. Treatment of the apo(a) KV-PD with O-glycosidase did not significantly affect its ability to bind to plasmin-treated fibrinogen. In addition, apo(a) KV-PD competed for the binding of ¹²⁵I-Lp(a) to plasmin-treated fibrinogen. An IC₅₀ of 7.90 ± 0.95 μM was obtained. Our data suggest that the KV-PD of apo(a) shares binding sites on plasmin-treated fibrinogen with Lp(a) and also may participate in the interaction of the Lp(a) particle with plasmin-treated fibrinogen.

Keywords

Apo(a) - kringle - fibrinogen

Full Text

References

References
1 Loscalzo J. Lipoprotein(a). A unique risk factor for atherothrombotic disease. Arteriosclerosis 1990; 10 (Suppl. 05) 672-9.
2 Nordestgaard BG. The vascular endothelial barrier – selective retention of lipoproteins. Current Opinion In Lipidology 1996; 7 (Suppl. 05) 269-73.
3 Simionescu M, Simionescu N. Proatherosclerotic events: pathobiochemical changes occurring in the arterial wall before monocyte migration. Faseb Journal 1993; 7 (Suppl. 14) 1359-66.
4 Fless GM, Rolih CA, Scanu AM. Heterogeneity of human plasma lipoprotein (a). Isolation and characterization of the lipoprotein subspecies and their apoproteins. J Biol Chem 1984; 259 (Suppl. 18) 11470-8.
5 McLean JW. et al. cDNA sequence of human apolipoprotein(a) is homo-logous to plasminogen. Nature 1987; 330 6144 132-7.
6 Eaton DL. et al. Partial amino acid sequence of apolipoprotein(a) shows that it is homologous to plasminogen. Proceedings of the National Academy of Sciences of the USA 1987; 84 (Suppl. 10) 3224-8.
7 Lucas MA, Fretto LJ, McKee PA. The binding of human plasminogen to fibrin and fibrinogen. J Biol Chem 1983; 258 (Suppl. 07) 4249-56.
8 Wiman B, Wallen P. The specific interaction between plasminogen and fibrin. A physiological role of the lysine binding site in plasminogen. Thrombosis Research 1977; 10 (Suppl. 02) 213-22.
9 Whitaker AN. et al. The binding of glu- and lys-plasminogens to fibrin and their subsequent effects on fibrinolysis. Thrombosis Research 1980; 19 (Suppl. 03) 381-91.
10 Fleury V, Anglés-Cano E. Characterization of the Binding of Plasminogen to Fibrin Surfaces: The Role of Carboxy-Terminal Lysines. Biochemistry 1991; 30: 7630-8.
11 Loscalzo J. et al. Lipoprotein(a), fibrin binding, and plasminogen activation. Arteriosclerosis 1990; 10 (Suppl. 02) 240-5.
12 Miles L. et al. Interaction of Lipoprotein(a) with the Extracellular Matrix. Fibrinolysis and Proteolysis 1998; 12: 79-83.
13 Rouy D. et al. Apolipoprotein(a) and plasminogen interactions with fibrin: a study with recombinant apolipoprotein(a) and isolated plasminogen fragments. Biochemistry 1992; 31 (Suppl. 27) 6333-9.
14 Harpel PC, Gordon BR, Parker TS. Plasmin catalyzes binding of lipoprotein (a) to immobilized fibrinogen and fibrin. Proceedings of the National Academy of Sciences of the USA 1989; 86 (Suppl. 10) 3847-51.
15 Xue S. et al. Comparison of the effects of Apo(a) kringle IV-10 and plasminogen kringles on the interactions of lipoprotein(a) with regulatory molecules. Thromb Haemost 1999; 81 (Suppl. 03) 428-35.
16 Lou XJ. et al. Fibrinogen deficiency reduces vascular accumulation of apolipoprotein(a) and development of atherosclerosis in apolipoprotein(a) transgenic mice. Proc Natl Acad Sci USA 1998; 95 (Suppl. 21) 12591-5.
17 Lawn RM. et al. Atherogenesis in transgenic mice expressing human apolipoprotein(a). Nature 1992; 360 6405 670-2.
18 Palabrica TM. et al. Antifibrinolytic activity of apolipoprotein(a) in vivo: human apolipoprotein(a) transgenic mice are resistant to tissue plasminogen activator-mediated thrombolysis. Nature Medicine 1995; 1 (Suppl. 03) 256-9.
19 Huby T. et al. Characterization of the N-terminal and C-terminal domains of human apolipoprotein(a): relevance to fibrin binding. Biochemistry 1995; 34 (Suppl. 22) 7385-93.
20 LoGrasso PV, Cornell KS, Boettcher BR. Cloning, expression, and characterization of human apolipoprotein(a) kringle IV37. J Biol Chem 1994; 269 (Suppl. 34) 21820-7.
21 Klezovitch O, Scanu AM. Lys and fibrinogen binding of wild-type (Trp72) and mutant (Arg72) human apo(a) kringle IV-10 expressed in E coli and CHO cells. Arterioscler Thromb Vasc Biol 1996; 16 (Suppl. 03) 392-8.
22 Scanu AM. et al. Rhesus monkey lipoprotein(a) binds to lysine Sepharose and U937 monocytoid cells less efficiently than human lipoprotein(a). Evidence for the dominant role of kringle 4 (37). J Clin Invest 1993; 91 (Suppl. 01) 283-91.
23 Scanu AM. et al. A single point mutation (Trp72—>Arg) in human apo(a) kringle 4-37 associated with a lysine binding defect in Lp(a). Biochimica et Biophysica Acta 1994; 1227 1-2 41-5.
24 Klezovitch O, Edelstein C, Scanu AM. Evidence that the fibrinogen binding domain of Apo(a) is outside the lysine binding site of kringle IV-10: a study involving naturally occurring lysine binding defective lipoprotein(a) phenotypes. J Clin Invest 1996; 98 (Suppl. 01) 185-91.
25 Ernst A. et al. Identification of two functionally distinct lysine-binding sites in kringle 37 and in kringles 32-36 of human apolipoprotein(a). J Biol Chem 1995; 270 (Suppl. 11) 6227-34.
26 Chenivesse X. et al. Expression of a recombinant kringle V of human apolipoprotein(a): antibody characterization and species specificity. Protein Expression & Purification 1996; 8 (Suppl. 02) 145-50.
27 Chang Y. et al. Structure and ligand binding determinants of the recombinant kringle 5 domain of human plasminogen. Biochemistry 1998; 37 (Suppl. 10) 3258-71.
28 McCance SG, Menhart N, Castellino FJ. Amino acid residues of the kringle-4 and kringle-5 domains of human plasminogen that stabilize their interactions with omega-amino acid ligands. J Biol Chem 1994; 269 (Suppl. 51) 32405-10.
29 Sanger F, Nicklen S, Coulson AR. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci USA 1977; 74 (Suppl. 12) 5463-7.
30 Zhang X-P, Kamata T, Yokoyama K, Puzon-McLaughlin W, Takada Y. Specific interaction of the recombinant disintegrin-like domain of MDC-15 (Metargidin, ADAM-15) with integrin αvβ3. J Biol Chem 2001; 273 (Suppl. 13) 7345-50.
31 Deutsch DG, Mertz ET. Plasminogen: purification from human plasma by affinity chromatography. Science 1970; 170 (Suppl. 962) 1095-6.
32 Miles LA, Dahlberg CM, Plow EF. The cell-binding domains of plasminogen and their function in plasma. J Biol Chem 1988; 263 (Suppl. 24) 11928-34.
33 Shepherd J, Bedford DK, Morgan HG. Radioiodination of human low density lipoprotein: a comparison of four methods. Clinica Chimica Acta 1976; 66 (Suppl. 01) 97-109.
34 Miles LA. et al. Interaction of Lp(a) with plasminogen binding sites on cells. Thromb Haemost 1995; 73 (Suppl. 03) 458-65.
35 Fraker PJ, Speck Jr JC. Protein and cell membrane iodinations with a sparingly soluble chloroamide, 1,3,4,6-tetrachloro-3a,6a-diphrenylglycoluril. Biochem Biophys Res Commun 1978; 80 (Suppl. 04) 849-57.
36 Scanu AM, Edelstein C. Kringle-dependent structural and functional polymorphism of apolipoprotein (a). Biochimica et Biophysica Acta 1995; 1256 (Suppl. 01) 1-12.
37 Bonen DK. et al. Expression of a recombinant apolipoprotein(a) in HepG2 cells. Evidence for intracellular assembly of lipoprotein(a). J Biol Chem 1997; 272 (Suppl. 09) 5659-67.
38 Sangrar W, Marcovina SM, Koschinsky ML. Expression and characterization of apolipoprotein(a) kringle IV types 1, 2 and 10 in mammalian cells. Protein Engineering 1994; 7 (Suppl. 05) 723-31.
39 Makino K, Furbee JW, Scanu AM, Fless GM. Effect of glycation on the properties of lipoprotein(a). Arterioscler Thromb Vasc Biol 1995; 15 (Suppl. 03) 385-91.
40 Hughes SD. et al. Lipoprotein(a) vascular accumulation in mice. In vivo analysis of the role of lysine binding sites using recombinant adenovirus. J Clin Invest 1997; 100 (Suppl. 06) 1493-500.