Affinity chromatography of thrombin on modified polystyrene resins

Area of Science:

• Biochemistry
• Polymer Chemistry
• Chromatography

Background:

• Antithrombin III is a key regulator of blood coagulation.
• Thrombin is a crucial enzyme in the coagulation cascade.
• Affinity chromatography is a powerful technique for protein purification.

Purpose of the Study:

• To develop and characterize novel insoluble polystyrenes for specific thrombin affinity.
• To investigate the influence of polymer characteristics on thrombin purification performance.
• To achieve high-purity, biologically active thrombin using the developed affinity resins.

Main Methods:

• Synthesis of insoluble polystyrenes substituted with sulphonate and L-arginyl methyl ester (PAOM).
• Characterization of polymer properties: substitution ratio, particle size, and affinity constant.
• Affinity liquid chromatography using the PAOM resins for thrombin purification.
• Activation of human prothrombin complex concentrate and application onto the chromatography gel.
• Analysis of purified thrombin for specific activity and recovery of biological activity.

Main Results:

• PAOM resins exhibit specific, reversible binding affinity for thrombin.
• Purified thrombin retains high biological activity after elution.
• Optimization of polymer characteristics (substitution ratio, particle size, affinity constant) enhances purification efficiency.
• High recovery of biologically active human thrombin was achieved from activated prothrombin complex concentrate.

Conclusions:

• PAOM-substituted polystyrenes are effective stationary phases for affinity chromatography of thrombin.
• The developed resins allow for the purification of highly active thrombin with high recovery.
• This method provides a valuable tool for obtaining pure, functional thrombin for research and therapeutic applications.