Neutron and X-ray crystallographic analysis of the human α-thrombin-bivalirudin complex at pD 5.0: protonation states and hydration structure of the enzyme-product complex.
Research article published in Biochimica et biophysica acta (2013)
Abstract
The protonation states and hydration structures of the α-thrombin-bivalirudin complex were studied by joint XN refinement of the single crystal X-ray and neutron diffraction data at resolutions of 1.6 and 2.8Å, respectively. The atomic distances were estimated by carrying out X-ray crystallographic analysis at 1.25Å resolution. The complex represents a model of the enzyme-product (EP) complex of α-thrombin. The neutron scattering length maps around the active site suggest that the side chain of H57/H was deuterated. The joint XN refinement showed that occupancies for Dδ1 and Dε2 of H57/H were 1.0 and 0.7, respectively. However, no significant neutron scattering length density was observed around the hydroxyl oxygen Oγ of S195/H, which was close to the carboxylic carbon atom of dFPR-COOH. These observations suggest that the Oγ atom of S195/H is deprotonated and maintains its nucleophilicity in the EP complex. In addition to the active site, the hydration structures of the S1 subsite and the Exosite I, which are involved in the recognition of bivalirudin, are presented.
Abstract sourced from PubMed (NCBI) for the cited record. See the original publication for the authoritative version.
Summary
The protonation states and hydration structures of the α-thrombin-bivalirudin complex were studied by joint XN refinement of the single crystal X-ray and neutron diffraction data at resolutions of 1.6 and 2.8Å, respectively. The atomic distances were estimated by carrying out X-ray crystallographic...
Why This Matters for Hirudotherapy
This study employed joint X-ray and neutron crystallography to resolve the protonation states and hydration structure of the human α-thrombin–bivalirudin enzyme-product complex at near-atomic resolution. Key findings include the deuteration state of the catalytic histidine (H57), apparent deprotonation of the serine Oγ nucleophile (S195), and detailed hydration at the S1 subsite and Exosite I — both critical for bivalirudin recognition. For ASH, this provides atomic-level structural insight into how a hirudin-derivative engages thrombin's active site and exosites, deepening mechanistic understanding of leech-inspired anticoagulation. CAVEAT: This is a purely in-vitro biophysical study of the synthetic pharmaceutical bivalirudin; it does not involve leeches, whole saliva, or in-vivo therapeutic application, and its relevance is limited to molecular pharmacology of a hirudin analog.
Citation
Neutron and X-ray crystallographic analysis of the human α-thrombin-bivalirudin complex at pD 5.0: protonation states and hydration structure of the enzyme-product complex.
Yamada et al. · Biochimica et biophysica acta, 2013
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