PDBsum entry 6v5t

Hydrolase

PDB id: 6v5t

Contents

Protein chain

290 a.a.

Ligands

GOL

SO4

Waters ×103

References listed in PDB file

Key reference

• Title: ¹⁹f nmr reveals the conformational properties of free thrombin and its zymogen precursor prethrombin-2.
• Authors: E.A.Ruben, P.S.Gandhi, Z.Chen, S.K.Koester, G.T.Dekoster, C.Frieden, E.Di cera.
• Ref.: J Biol Chem, 2020, 295, 8227-8235. [DOI no: 10.1074/jbc.RA120.013419]
• PubMed id: 32358061

Abstract

The conformational properties of trypsin-like proteases and their zymogen forms remain controversial because of a lack of sufficient information on their free forms. Specifically, it is unclear whether the free protease is zymogen-like and shifts to its mature form upon a ligand-induced fit or exists in multiple conformations in equilibrium from which the ligand selects the optimal fit via conformational selection. Here we report the results of ¹⁹F NMR measurements that reveal the conformational properties of a protease and its zymogen precursor in the free form. Using the trypsin-like, clotting protease thrombin as a relevant model system, we show that its conformation is quite different from that of its direct zymogen precursor prethrombin-2 and more similar to that of its fully active Na⁺-bound form. The results cast doubts on recent hypotheses that free thrombin is zymogen-like and transitions to protease-like forms upon ligand binding. Rather, they validate the scenario emerged from previous findings of X-ray crystallography and rapid kinetics supporting a pre-existing equilibrium between open (E) and closed (E*) forms of the active site. In this scenario, prethrombin-2 is more dynamic and exists predominantly in the E* form, whereas thrombin is more rigid and exists predominantly in the E form. Ligand binding to thrombin takes place exclusively in the E form without significant changes in the overall conformation. In summary, these results disclose the structural architecture of the free forms of thrombin and prethrombin-2, consistent with an E*-E equilibrium and providing no evidence that free thrombin is zymogen-like.