title: Inactivation of human coagulation factor X by a protease of the pathogen Capnocytophaga canimorsus.
abstract: Essentials Capnocytophaga canimorsus causes severe dog bite related blood stream infections. We investigated if C. canimorsus contributes to bleeding abnormalities during infection. The C. canimorsus protease CcDPP7 causes factor X dysfunction by N-terminal cleavage. CcDPP7 inhibits coagulation in vivo, which could promote immune evasion and trigger hemorrhage.
  
  
  
  Background Capnocytophaga canimorsus is a Gram-negative bacterium that is present in the oral flora of dogs and causes fulminant sepsis in humans who have been bitten, licked, or scratched. In patients, bleeding abnormalities, such as petechiae, purpura fulminans, or disseminated intravascular coagulation (DIC), occur frequently. Objective To investigate whether C. canimorsus could actively contribute to these bleeding abnormalities. Methods Calibrated automated thrombogram and clotting time assays were performed to assess the anticoagulant activity of C. canimorsus 5 (Cc5), a strain isolated from a fatal human infection. Clotting factor activities were measured with factor-deficient plasma. Factor X cleavage was monitored with the radiolabeled zymogen and western blotting. Mutagenesis of Cc5 genes encoding putative serine proteases was performed to identify the protease that cleaves FX. Protein purification was performed with affinity chromatography. Edman degradation allowed the detection of N-terminal cleavage of FX. Tail bleeding times were measured in mice. Results We found that Cc5 inhibited thrombin generation and increased the prothrombin time and the activated partial thromboplastin time of human plasma via FX cleavage. A mutant that was unable to synthesize a type 7 dipeptidyl peptidase (DPP7) of the S46 serine protease family failed to proteolyse FX. The purified protease (CcDPP7) cleaved FX heavy and light chains from the N-terminus, and was active in vivo after intravenous injection. Conclusions This is, to our knowledge, the first study demonstrating a detailed mechanism for FX inactivation by a bacterial protease, and it is the first functional study associating DPP7 proteases with a potentially pathogenic outcome.
authors.affiliation: Department of Biology, University of Namur, Namur, Belgium.
authors.forename: K
authors.initials: K
authors.lastname: Hack
authors.affiliation: Department of Biology, University of Namur, Namur, Belgium.
authors.forename: F
authors.initials: F
authors.lastname: Renzi
authors.affiliation: Department of Biology, University of Namur, Namur, Belgium.
authors.forename: E
authors.initials: E
authors.lastname: Hess
authors.affiliation: Department of Biology, University of Namur, Namur, Belgium.
authors.forename: F
authors.initials: F
authors.lastname: Lauber
authors.affiliation: Department of Pharmacy, University of Namur, Namur, Belgium.
authors.forename: J
authors.initials: J
authors.lastname: Douxfils
authors.affiliation: Department of Pharmacy, University of Namur, Namur, Belgium.
authors.forename: J M
authors.initials: JM
authors.lastname: Dogné
authors.affiliation: Department of Biology, University of Namur, Namur, Belgium.
authors.forename: G R
authors.initials: GR
authors.lastname: Cornelis
mesh_terms: D000818:Animals; D001733:Bites and Stings / Q000382:microbiology*; D002206:Capnocytophaga / Q000201:enzymology*; D002384:Catalysis; D004211:Disseminated Intravascular Coagulation / Q000382:microbiology*; D005170:Factor X / Q000037:antagonists & inhibitors*; D064368:Healthy Volunteers; D006801:Humans; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D009154:Mutation; D010314:Partial Thromboplastin Time; D010447:Peptide Hydrolases / Q000737:chemistry*; D010957:Plasmids / Q000378:metabolism; D000072417:Protein Domains; D018805:Sepsis / Q000382:microbiology; D017422:Sequence Analysis, DNA
chemical_list: D005170:Factor X; D010447:Peptide Hydrolases
keywords: bacterial infection; coagulation; factor X; protease; sepsis