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Thromb Haemost 2008; 100(05): 766-772
DOI: 10.1160/TH08-02-0106
Blood Coagulation, Fibrinolysis and Cellular Haemostasis
Schattauer GmbH

Modelling and expression studies of two novel mutations causing factor V deficiency

Daniel Delev
1   Institute of Experimental Haematology and Transfusion Medicine, University Clinic Bonn, Bonn, Germany
,
Anna Pavlova
1   Institute of Experimental Haematology and Transfusion Medicine, University Clinic Bonn, Bonn, Germany
,
Stefan Heinz
2   Institute for Transfusion Medicine and Immunohaematology, Red Cross Blood Donor Service, Baden-Wuerttemberg-Hessen, University Clinic Frankfurt, Germany
,
Mathias Costa Blaise
3   Department of Pharmacology and Experimental Neuroscience, University of Nebraska Medical Center, Omaha, Nebraska, USA
,
Tamir Chandra
4   Cancer Research UK, Cambridge Research Institute, Li Ka Shing Centre, Cambridge, UK
,
Bernd Poetsch
1   Institute of Experimental Haematology and Transfusion Medicine, University Clinic Bonn, Bonn, Germany
,
Erhard Seifried
2   Institute for Transfusion Medicine and Immunohaematology, Red Cross Blood Donor Service, Baden-Wuerttemberg-Hessen, University Clinic Frankfurt, Germany
,
Johannes Oldenburg
1   Institute of Experimental Haematology and Transfusion Medicine, University Clinic Bonn, Bonn, Germany
› Author AffiliationsFinancial support: The work was supported by grants from the National Genome Research Net Cardiovascular Diseases (BMBF/DLR-01GS0424/NHK-S12T21).
Further Information

Publication History

Received 24 February 2008

Accepted after major revision 12 August 2008

Publication Date:
22 November 2017 (online)

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Summary

Human coagulation factor V (FV), a non-enzymatic cofactor of the prothrombinase complex, is required for the rapid generation of thrombin. FV deficiency is a rare autosomal recessive bleeding disorder. We describe two novel mutations,Tyr91Asn and Asp2098Tyr,found in two probands with a residual FV activity of 51% and 4%, respectively. Modelling and structural analysis of these mutations were performed following short-duration molecular dynamics (MD) simulation.Asp2098Tyr lead to abolishment of the highly conserved salt bridge Asp2098-Arg2171 presumably required for structural integrity of the C2 domain. MD studies suggest that additional conformational changes resulting from this mutation involve local rearrangements at Tyr2063 and Tyr2064 and so affect the phospholipid-membrane binding. MD modelling of the Try91Asn mutant revealed a conformational change nearby the Cu2+ binding site that could affect overall stabilization of the heavy and light chains. These findings suggest that both mutations influence the structural integrity of FV protein. Transient expression data of wild-type and mutant FV variants in 293T human embryonic kidney cells showed FV-specific activity reduced to 26% for Asp2098Tyr and 56% for Tyr91Asn compared to that of wild-type. Thus, both the data from the short duration molecular dynamic simulation and from expression analysis indicate alterations of the FV protein variants that explain the clinical phenotype.

Keywords

Factor V deficiency - novel mutations - MD simulation - transient expression studies
 

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