Small Molecule Probes That Perturb A Protein-protein Interface In Antithrombin

Area of Science:

• Medicinal Chemistry
• Biochemistry
• Structural Biology

Background:

• Small molecule probes can perturb protein-protein interactions (PPIs) by inducing structural changes.
• Alpha-antithrombin, a serpin, exists in a metastable oligomeric state relevant to disease.
• Serpinopathies are associated with the formation of oligomeric serpin fibrils.

Purpose of the Study:

• To design and synthesize small molecule probes targeting the alpha-antithrombin dimer interface.
• To investigate the effect of these probes on alpha-antithrombin oligomerization.
• To explore the potential of these molecules in modulating serpin behavior relevant to disease.

Main Methods:

• Application of the Exploring Key Orientations (EKO) strategy for probe design.
• Synthesis of piperidinone-piperidine chemotype derivatives.
• Analysis of protein oligomerization using non-denaturing polyacrylamide gel electrophoresis (PAGE).
• Assay of alpha-antithrombin's inhibitor activity in a thrombin-catalyzed reaction.
• Kinetic analysis of probe-induced oligomerization.
• Molecular modeling experiments.

Main Results:

• Several synthesized derivatives of compound 1 significantly accelerated the oligomerization of monomeric alpha-antithrombin.
• Probe-induced loss of alpha-antithrombin's inhibitor activity was observed.
• Probes featuring O-benzyl-protected serine side-chains demonstrated the highest catalytic activity.
• Modeling experiments provided proposed reasons for the observed activity differences.

Conclusions:

• This study presents one of the first examples of small molecules designed to target a protein-protein interface involved in serpin oligomerization.
• The findings highlight the potential of small molecules to modulate alpha-antithrombin aggregation.
• The research has implications for understanding and potentially treating serpinopathies.