Thrombin inhibitor design: X-ray and solution studies provide a novel P1 determinant

Area of Science:

• Biochemistry
• Structural Biology
• Pharmacology

Background:

• Thrombin is a key enzyme in hemostasis.
• Understanding ligand interactions at the active site is crucial for developing inhibitors.
• Previous studies hinted at the role of ions in enzyme activity.

Purpose of the Study:

• To elucidate the binding modes of proflavin and 6-fluorotryptamine to thrombin.
• To investigate the influence of a sodium ion on ligand binding.
• To inform the design of novel thrombin inhibitors.

Main Methods:

• X-ray crystallography to determine ligand-thrombin complex structures.
• Direct proflavin titrations.
• Chromogenic activity assays measuring thrombin activity at varying sodium concentrations.

Main Results:

• Confirmed proflavin binding in the S1 pocket.
• Identified a novel binding mode for 6-fluorotryptamine at the thrombin active site.
• Demonstrated a functional linkage between a sodium ion binding site and the S1 pocket.

Conclusions:

• The crystal structures provide detailed insights into thrombin-ligand interactions.
• The sodium ion likely influences the thrombin S1 pocket conformation or ligand binding.
• The unique 6-fluorotryptamine binding mode offers a template for designing neutral thrombin inhibitors.