Soluble phospholipids enhance factor Xa-catalyzed prothrombin activation in solution

Area of Science:

• Biochemistry
• Molecular Biology
• Hematology

Background:

• Acidic phospholipids are crucial for prothrombin activation, a key step in blood coagulation.
• The precise mechanisms by which phospholipids influence this process remain incompletely understood.

Purpose of the Study:

• To investigate the effects of short-chain acidic phospholipids, phosphatidylserine (C6PS), phosphatidylglycerol (C6PG), and phosphatidylcholine (C6PC), on prothrombin activation by factor Xa.
• To elucidate the role of specific lipid-protein interactions in regulating coagulation factor activity.

Main Methods:

• Kinetic studies of prothrombin activation and synthetic substrate hydrolysis by factor Xa in the presence of various short-chain phospholipids.
• Determination of critical micellar concentrations (CMCs) for the studied lipids.
• Analysis of prothrombin thermal denaturation profiles.

Main Results:

• Short-chain phosphatidylserine (C6PS) enhanced human prothrombin activation by human factor Xa in a calcium-dependent manner (up to 60-fold).
• C6PS inhibited factor Xa substrate hydrolysis, suggesting specific binding to factor Xa.
• Phosphatidylglycerol (C6PG) and phosphatidylcholine (C6PC) showed minimal effects on prothrombin activation and substrate hydrolysis.
• Both C6PS and C6PG altered prothrombin thermal denaturation in a calcium-independent, dose-dependent manner.

Conclusions:

• Specific binding sites for phosphatidylserine (PS) on factor Xa (Kd ~73 µM) regulate its serine protease activity.
• PS binding is a key regulator of prothrombin activation rate.
• Potential weaker, calcium-independent binding sites on prothrombin may also influence activation through conformational changes.