Studies on the Effect of Calcium in Interactions Between Heparin and Heparin Cofactor II Using Surface Plasmon Resonance

Abstract

Heparin is the most acidic polysaccharide in the human body and as a result interacts with many cationic species, including ions and proteins, giving rise to myriad biologic activities. Heparin cofactor II (HCII) is a serine protease inhibitor that resembles antithrombin (ATIII) in its ability to be activated by heparin. The interaction of heparin with HCII has been the focus of many studies using affinity chromatography and fluorescence spectroscopy. In this study, surface plasmon resonance (SPR) spectroscopy was used to quantitatively measure the interaction of heparin and HCII using a heparin biochip prepared by covalently immobilizing preformed albumin-heparin conjugate. HCII contains multiple EF hand domains that represent putative calcium ion binding sites. The interactions of HCII with heparin, low-molecular-weight heparin, and heparin oligosaccharides (disaccharide, tetrasaccharide, hexasaccharide) were examined in solution competition experiments using SPR. The results also showed while calcium ions enhanced the heparin/HCII interaction, the activity of heparin-HCII complex against thrombin was not calcium dependent but can be enhanced by the presence of calcium.

Keywords

Heparin Heparin cofactor II Interaction Calcium Surface plasmon resonance

References

Capila I , Linhardt RJ . Heparin-protein interactions. Angew Chemie Int Ed 2002;41:390.

Bernfield M , Gotte M , Park PW , Reizes O , Fitzgerald ML , Lincecum J , Zako M. Functions of cell surface heparan sulfate proteoglycans. Annu Rev Biochem 1999;68:729.

Tollefsen DM . Insight into the mechanism of action of heparin cofactor II. Thromb Haemost 1995;74:1209.

Liaw PC , Austin RC , Fredenburgh JC , Stafford AR , Weitz JI . Comparison of heparin- and dermatan sulfate-mediated catalysis of thrombin inactivation by heparin cofactor II. J Biol Chem 1999;274:27597.

Rich RL , Myszka DG . Advances in surface plasmon resonance biosensor analysis. Curr Opin Biotechnol 2000;11:54.

Capila I , Hernaiz MJ , Mo YD , Mealy TR , Campos B , Linhardt RJ , Seaton BA . Annexin V-heparin oligosaccharide complex suggests heparan sulfate-mediated assembly on cell surfaces. Structure 2001;9:57.

Dong J , Peters-Libeu C , Weisgraber KH , et al. Interaction of the N-terminal domain of apolipoprotein E4 with heparin. Biochemistry 2001;40:2826.

Marks RM , Lu H , Sundaresan R , et al. Probing the interaction of dengue virus envelope protein with heparin: Assessment of glycosaminoglycan-derived inhibitors. J Med Chem 2001;44:2178.

Kamei K , Wu X , Xu X , et al. The analysis of heparin-protein interactions using evanescent wave biosensor with regioselectively desulfated heparins as the ligands. Anal Biochem 2001;295:203.

10.

Hayakawa Y , Hayashi T , Lee JB , Ozawa T , Sakuragawa N. Activation of heparin cofactor II by calcium spirulan. J Biol Chem 2000;275(15):11379.

11.

Zhang F , Fath M , Marks R , Linhardt RJ . A highly stable covalent conjugated heparin biochip for heparin-protein interactions studies. Anal Biochem 2002;304:271.

12.

Walenga JM , Fareed J , Hoppensteadt D , Emanuele RM . In vitro evaluation of heparin fractions: Old vs. new methods. Crit Rev Clin Lab Sci 1986;22 (4):361.

13.

Kim YS , Linhardt RJ . Structural features of heparin and their effect on heparin cofactor II mediated inhibition of thrombin. Thromb Res 1989;53:55.

14.

Liu D , Shriver Z , Godavarti R , Venkataraman G , Sasisekharan R. The calcium-binding sites of heparinase I from Flavobacterium heparinum are essential for enzymatic activity. J Biol Chem 1999;274:4089.

15.

Tollefsen DM . The interaction of glycosaminoglycans with heparin cofactor II. Ann NY Acad Sci 1994;714:21.

16.

Lweit-Bently A , Rety S. EF-hand calcium binding proteins. Curr Opin Struct Biol 2000;10:637.

17.

Grant D , Moffat CF , Long WF , Williamson FB . Optical rotation changes in chemically modified heparins as a guide to anionic groups involved in Ca2+ binding. Biochem Soc Trans 1991;19:393S.

18.

Grant D , Moffat CF , Long WF , Williamson FB . Ca (2+)-heparin interaction investigated polarimetrically. Biochem Soc Tran 1991;19:391S.

19.

Grant D , Long WF , Williamson FB . NMR spectroscopy of Ca (2+)-heparin suggests delocalized binding of the cation. Biochem Soc Trans 1991;19:390S.

20.

Kluszynski BA , Kim C , Faulk WP . Zinc as a cofactor for heparin neutralization by histidine-rich glycoprotein. J Biol Chem 1997;272:13541.

21.

Baglin TP , Carell RW , Church FC , Esmon CT , Huntington JA . Crystal structures of native and thrombin-complexed heparin cofactor II reveal a multistep allosteric mechanism. Proc Natl Acad Sci USA 2002;99:11079.