Enrico Di Cera, M.D.
Alice A. Doisy Professor & Chair
Structural enzymology of coagulation factors.
Research Interests
Our research builds upon 30+ years of continuously funded work on the enzymology and structural biology of proteins involved in blood coagulation and on mechanisms of ligand binding.
We are interested in the structural enzymology of proteins responsible for blood coagulation, with the goal of unraveling their molecular architecture and mechanisms. Our approach uses a combination of conventional (rapid kinetics, protein engineering, X-ray crystallography) and innovative (smFRET, cryo-EM) biophysical techniques.
We currently focus on the structure, function and regulation of prothrombin (HL049413), protein C (HL139554) and factor V (HL147821). Also of interest is the molecular basis of protease specificity.
Research Highlights
Cryo-EM structures of human coagulation factors V and Va. Ruben EA, Rau MJ, Fitzpatrick JAJ, Di Cera E. Blood 137(22):3137, 2021.
Cryo-EM structure of the prothrombin-prothrombinase complex. Ruben EA, Summers B, Rau MJ, Fitzpatrick JAJ, Di Cera E. Blood 139(24):3463, 2022.
Cryo-EM structure of coagulation factor Va bound to activated protein C. Mohammed BM, Basore K, Di Cera E. Blood 145(26):3166, 2025.
Recent Publications
Structural determinants of protease-activated receptor 1 cleavage by activated protein C
Structural determinants of protease-activated receptor 1 cleavage by activated protein C
Activated protein C (APC) performs cytoprotective functions mediated by cleavage of the protease-activated receptor 1 (PAR1) in the presence of the endothelial protein C receptor and signaling through β-arrestin-2. APC cleaves PAR1 at R41 and R46, but the specificity of the reaction is low. In contrast, thrombin cleaves PAR1 at R41 only in a reaction that is independent of endothelial protein C receptor, producing a proinflammatory response mediated by signaling through G-protein intermediates and features high specificity. The molecular basis of this difference between APC and thrombin remains unknown.
Cryo-EM structure of coagulation factor Va bound to activated protein C
Cryo-EM structure of coagulation factor Va bound to activated protein C
Coagulation factor Va (FVa) is the cofactor component of the prothrombinase complex required for rapid generation of thrombin from prothrombin in the penultimate step of the coagulation cascade. In addition, FVa is a target for proteolytic inactivation by activated protein C (APC). Like other protein-protein interactions in the coagulation cascade, the FVa-APC interaction has long posed a challenge to structural biology and its molecular underpinnings remain unknown. A recent cryogenic electron microscopy (cryo-EM) structure of FVa has revealed the arrangement of its A1-A2-A3-C1-C2 domains and the environment of the sites of APC cleavage at R306 and R506. Here, we report the cryo-EM structure of the FVa-APC complex at 3.15 Å resolution in which the protease domain of APC engages R506 in the A2 domain of FVa through electrostatic interactions between positively charged residues in the 30-loop and 70-loop of APC and an electronegative surface of FVa. The auxiliary γ-carboxyglutamic acid and epidermal growth factor domains of APC are highly dynamic and point to solvent, without making contacts with FVa. Binding of APC displaces a large portion of the A2 domain of FVa and projects the 654VKCIPDDDEDSYEIFEP670 segment as a “latch,” or exosite ligand, over the 70-loop of the enzyme. The latch induces a large conformational change of the autolysis loop of APC, which in turn promotes docking of R506 into the primary specificity pocket. The cryo-EM structure of the FVa-APC complex validates the bulk of existing biochemical data and offers molecular context for a key regulatory interaction of the coagulation cascade.
Replacement of a single residue changes the primary specificity of thrombin
Replacement of a single residue changes the primary specificity of thrombin
Thrombin prefers substrates carrying Arg at the site of cleavage (P1) because of the presence of D189 in the primary specificity (S1) pocket but can also cleave substrates carrying Phe at P1. The structural basis of this property is unknown.
Another cryo-EM success: structure of FXIII
Safety and tolerability of the protein C activator AB002 in end-stage renal disease patients on hemodialysis: a randomized phase 2 trial
Safety and tolerability of the protein C activator AB002 in end-stage renal disease patients on hemodialysis: a randomized phase 2 trial
The protein C system regulates blood coagulation, inflammation, and vascular integrity. AB002 is an injectable protein C activating enzyme under investigation to safely prevent and treat thrombosis. In preclinical models, AB002 is antithrombotic, cytoprotective, and anti-inflammatory. Since prophylactic use of heparin is contraindicated during hemodialysis in some end-stage renal disease (ESRD) patients, we propose using AB002 as a short-acting alternative to safely limit blood loss due to clotting in the dialysis circuit.
