Kinetic and hydrodynamic analysis of blood clotting factor V-membrane binding

Marc Lee Pusey, Lawrence D. Mayer, G. Jason Wei, Victor A. Bloomfield, Gary L. Nelsestuen

Research output: Contribution to journalArticle

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Abstract

The kinetics and hydrodynamic properties of factor V-membrane interaction were characterized. Factor V bound to membranes containing acidic phospholipids with a high collisional efficiency. For membranes of 20% phosphatidylserine-80% phosphatidylcholine, an association rate constant of (1.13 ± 0.10) × 108 M-1 s-1 was obtained. These membranes contained about 20 factor V binding sites per vesicle of 3.6 × 106 daltons. This association rate represented about a 30% collisional efficiency. Dissociation of factor V was measured by a fluorescence energy transfer method with a dissociation rate constant of 0.0055 s-1 at 10°C. The equilibrium dissociation constant for binding to these membranes at 10°C and 0.14 M ionic strength was 5 × 10-11 M. Ionic strength, pH, calcium, and charge density in the membrane had large effects on the rate of factor V-membrane dissociation, indicating a strongly ionic interaction between protein and membrane. In contrast, the association rate was nearly insensitive to ionic strength. The membrane-binding properties were relatively unchanged after thrombin digestion of factor V or after long-term protein storage which resulted in loss of procoagulant activity. Other proteins of the prothrombinase reaction greatly decreased the rate of factor Va-membrane dissociation. At protein saturation, factor V increased the hydrodynamic radius of phospholipid vesicles by 11.4 nm. In contrast, factor Va increased the hydrodynamic vesicle radius by only about 5 nm. The mass of membrane-bound protein was comparable for both proteins.

Original languageEnglish (US)
Pages (from-to)5262-5269
Number of pages8
JournalBiochemistry
Volume21
Issue number21
StatePublished - 1982

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Blood Coagulation Factors
Factor V
Blood Coagulation
Hydrodynamics
Blood
Membranes
Kinetics
Factor Va
Osmolar Concentration
Ionic strength
Association reactions
Proteins
Phospholipids
Membrane Proteins
Rate constants
Phosphatidylserines
Energy Transfer
Thromboplastin
Phosphatidylcholines
Thrombin

Cite this

Pusey, M. L., Mayer, L. D., Wei, G. J., Bloomfield, V. A., & Nelsestuen, G. L. (1982). Kinetic and hydrodynamic analysis of blood clotting factor V-membrane binding. Biochemistry, 21(21), 5262-5269.

Kinetic and hydrodynamic analysis of blood clotting factor V-membrane binding. / Pusey, Marc Lee; Mayer, Lawrence D.; Wei, G. Jason; Bloomfield, Victor A.; Nelsestuen, Gary L.

In: Biochemistry, Vol. 21, No. 21, 1982, p. 5262-5269.

Research output: Contribution to journalArticle

Pusey, ML, Mayer, LD, Wei, GJ, Bloomfield, VA & Nelsestuen, GL 1982, 'Kinetic and hydrodynamic analysis of blood clotting factor V-membrane binding', Biochemistry, vol. 21, no. 21, pp. 5262-5269.
Pusey ML, Mayer LD, Wei GJ, Bloomfield VA, Nelsestuen GL. Kinetic and hydrodynamic analysis of blood clotting factor V-membrane binding. Biochemistry. 1982;21(21):5262-5269.
Pusey, Marc Lee ; Mayer, Lawrence D. ; Wei, G. Jason ; Bloomfield, Victor A. ; Nelsestuen, Gary L. / Kinetic and hydrodynamic analysis of blood clotting factor V-membrane binding. In: Biochemistry. 1982 ; Vol. 21, No. 21. pp. 5262-5269.
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