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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
| Localization: |
Plasma |
| Mode of action: |
Enzyme component of the prothrombinase complex |
| Molecular weight: |
46,000 (human) (4)
45,300 (bovine) (5) |
| Extinction coefficient: |
| E |
|
= 11.6 (human) (6) |
|
|
= 12.4 (bovine) (7) |
|
| Specific activity: |
approximately 1000 units/mg |
| Structure: |
two subunits,
Mr=16,200 and 29,000 (human) (6), Mr=16,500 and 28,800 (bovine) (5),
NH2-terminal gla domain, two EGF domains |
| Percent carbohydrate: |
3.0 % (human) (8)
2.1 % (bovine) (8) |
| Post-translational modifications: |
eleven gla residues,
one
β-hydroxyaspartate |
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
Glyercol
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 |
|
U.S.
Pricing |
Product
inquiry |
SAMPLE
DATA SHEET |
ORDER
NOW! |
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).
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