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Coagulation Factor Xa

PARTICIPATION OF FACTOR Xa IN PROTHROMBINASE The participation of the serine protease factor Xa in the prothrombinase complex is illustrated. Membrane bound factor Xa binds to membrane bound factor Va to form the prothrombinase complex. This complex effectively converts the zymogen prothrombin (II) to the active serine protease thrombin (IIa) by proteolytic removal of the fragment 1.2 (F1.2) portion of prothrombin.

PURCHASING AND PRODUCT INFORMATION

Catalog Number HCXA-0060 HCBXA-0061 HCXA-GD HCXA-EGR HCXA-DEGR HCXA-BEGR BCXA-1060 BCXA-EGR BCXA-DEGR MCXA-5060*	Description Human Factor Xa Human β-Factor Xa Human Gla-Domainless β-Factor Xa Human Factor Xa -EGR Human Factor Xa - DEGR Human Factor Xa - BEGR (Biotin- EGR) Bovine Factor Xa Bovine Factor Xa - EGR Bovine Factor Xa - DEGR Mouse Factor Xa	Size 100 µg 100 µg 100 µg 100 µg 100 µg 100 µg 100 µg 100 µg 100 µg 50 µg	Formulation 50% glycerol/water (v/v) 50% glycerol/water (v/v) 10 mM Hepes, 50 mM NaCl, pH 7.4 20 mM Hepes, 150 mM NaCl, pH 7.4 20 mM Hepes, 150 mM NaCl, pH 7.4 20 mM Hepes, 150 mM NaCl, pH 7.4 50% glycerol/water (v/v) 20 mM Hepes, 150 mM NaCl, pH 7.4 20 mM Hepes, 150 mM NaCl, pH 7.4 50% glycerol/water (v/v)
Storage Glycerol formulation: -20oC Hepes formulation: -80oC	Purity >95% by SDS-PAGE	Activity Determination Factor X clotting or chromogenic assay	Shelf Life (properly stored) 12 months
Compliment fluorogenic substrate(s): HTI Catalog # SN-7		DOWNLOAD FACTOR XA PDF
		Sample Gel Information: Gel: Novex 4-12% Bis-Tris Load: Human Factor Xa, 1 µg per lane Buffer: MOPS Standard: SeeBluePlus 2; Myosin (191 kDa), Phosphorylase B (97 kDa), BSA (64 kDa), Glutamic Dehydrogenase (51 kDa), Alcohol Dehydrogenase (39 kDa), Carbonic Anhydrase (28 kDa), Myoglobin Red (19 kDa), Lysozyme (14 kDa) *Heavy chain is a doublet due to the presence of up to 50% beta form. The conversion of alpha-Xa to beta-Xa occurs by autocleavage of alpha-Xa by alpha-Xa resulting in the loss of a COOH-terminal peptide.
Sample publications referencing our Factor Xa: Monteiro, S., Biochem. J. (2005) 387, 871–877. (Prothrombin Activation) Zhang, D., et al., Biochemistry. 2006 November 28; 45(47): 14175–14182. (Prothrombin Activation) Kamata, K., et al., Proc. Natl. Acad. Sci. USA, Vol. 95, pp. 6630–6635, June 1998 (Crystallization) Yang, Y., et al., J Immunol. 2006 December 1; 177(11): 8219–8225. (Used as capture in ELISA) This publication list is not all encompassing, and is only meant to provide limited examples of how Haematologic Technologies' products are used.  We encourage you to search the literature for other examples pertinent to your experimentation, and to contact us with any technical questions.
U.S. Pricing	Product inquiry	SAMPLE DATA SHEET	ORDER NOW!

*Due to mouse plasma quality issues our mouse proteins are not being sold until further notice.  At the present time we cannot give a completion date.  Thank you.

Overview of Factor Xa

Activation of the zymogen, factor X, by either the intrinsic or extrinsic factor Xase complexes produces the active serine protease factor Xa (1,2). The activation of factor X requires proteolytic cleavage of the heavy chain, resulting in the release of an activation glycopeptide. The heavy chain region in factor Xa contains the serine protease catalytic domain, while the light chain, as in the zymogen, contains the membrane binding domain.

 
Factor Xa participates in the prothrombinase complex, which catalyzes the rapid conversion of prothrombin to thrombin. Prothrombinase is an enzyme complex composed of factor Xa (enzyme) and factor Va (cofactor) assembled on a cellular surface in the presence of calcium ions. Although factor Xa can independently catalyze the activation of prothrombin, the rate at which this reaction occurs is increased nearly 300,000-fold with complete assembly of the prothrombinase complex. The clotting activity of factor Xa in vivo is terminated by either inactivation of the cofactor, factor Va, or by direct inhibition of factor Xa by inhibitors, such as ATIII, after disassembly of the prothrombinase complex. 

In recent years, molecular biologists have utilized factor Xa for site specific cleavage of fusion proteins expressed in bacteria (9-12). A factor Xa-sensitive site is incorporated between the recombinant protein of interest and peptides or proteins which facilitate purification and/or expression. The target protein is released from the expressed hybrid by cleavage with factor Xa. The factor Xa can then be easily removed by affinity chromatography. 

Factor Xa is prepared by activating purified factor X with the factor X activator isolated from Russell's viper venom. Factor Xa is purified from the activation mixture by chromatography over benzamidine-Sepharose followed by gel filtration (1,3). Several modified forms of factor Xa are also available including: A) active-site blocked factor Xa containing either the tripeptide chloromethyl ketone inhibitor EGRck, or the fluorescent inhibitor Dansyl-EGRck; and B) human Gla-domainless β-factor Xa. The enzyme is supplied in 50% (vol/vol) glycerol/H₂O and should be stored at -20°C. Purity is determined by SDS-PAGE analysis and activity is measured in a factor Xa clotting assay and/or chromogenic substrate assay. 

Cleavage of Fusion Proteins

In addition to its broad application in coagulation research factor Xa can be used for site specific cleavage of fusion proteins. A factor Xa sensitive site is incorporated between the recombinant protein of interest and peptides or proteins which facilitate purification and/or expression.  The target protein is released from the expressed hybrid by cleavage with factor Xa. Factor Xa can then be easily removed by affinity chromatography. Lot to lot consistency ensures reproducible results every time. For experiments involving cell cultures, please contact us to discuss custom, low endotoxin lots designated for cell culture use.

Properties of Factor Xa

References 

1. Jesty, J., et al., Methods Enzymol., 45, 95 (1976). 
2. Jackson, C.M., Ann. Rev. Biochem., 49, 765 (1980). 
3. Krishnaswamy, S., et al., J.Biol. Chem., 262, 3291 (1987). 
4. Discipio, R.G., et al., Biochemistry, 16, 5253 (1977). 
5. Fujikawa, K. and Davie, E.W., Methods Enzymol., 45, 89 (1976). 
6. Krishnaswamy, S., et al., J. Biol. Chem., 261, 8997 (1986). 
7. Discipio, R.G., et al., Biochemistry, 16, 698 (1977). 
8. Jackson, C.M., et al., Biochemistry, 7, 4506 (1968). 
9. Fujikawa, K., Biochemistry, 13, 5290 (1974).
10. Nagai, K., et al., Proc. Natl. Acad. Sci. USA, 82, 7252 (1985). 
11. Aurell, L., et al., Thromb. Res., 11, 595 (1984). 
12. Nagai, K. and Thogersen, H., Nature, 309, 810 (1984).