Molecular Characterisation and Three-Dimensional Structural Analysis of Mutations in 21 Unrelated Families with Inherited Factor VII Deficiency
Flora Peyvandi
1
From the Haemophilia Centre and Haemostasis Unit, Department of Haematology, Royal Free and University College Medical School, Royal Free Campus, London, UK
2
Angelo Bianchi Bonomi Haemophilia and Thrombosis Center and Fondazione, Luigi Villa, IRCCS Maggiore Hospital and University of Milan, Italy
,
Vincent P. Jenkins
1
From the Haemophilia Centre and Haemostasis Unit, Department of Haematology, Royal Free and University College Medical School, Royal Free Campus, London, UK
,
Pier M. Mannucci
2
Angelo Bianchi Bonomi Haemophilia and Thrombosis Center and Fondazione, Luigi Villa, IRCCS Maggiore Hospital and University of Milan, Italy
,
Atto Billio
3
Department of Hematology, City Hospital, Bolzano, Italy
,
Sirous Zeinali
4
Pasteur Institute, Biotechnology Department, Tehran, Iran
,
Stephen J. Perkins
5
Department of Biochemistry and Molecular Biology, Royal Free and University College Medical School, Royal Free Campus, London, UK
,
David J. Perry
1
From the Haemophilia Centre and Haemostasis Unit, Department of Haematology, Royal Free and University College Medical School, Royal Free Campus, London, UK
› Author AffiliationsThe financial support of the Katharine Dormandy Trust for Haemophilia and Allied Disorders and the Angelo Bianchi Bonomi Foundation is gratefully acknowledged. We are also grateful to Dr. R. Sharifian, Dr. M. Lak and the Iranian Haemophilia Society for their help and assistance in the identification and collection of samples.
Factor VII (FVII) is a four-domain glycoprotein that plays a critical role in the initiation of blood coagulation. Hereditary deficiencies of this plasma protein results in a bleeding diathesis that varies in severity amongst affected patients. We have analysed the FVII gene in 27 patients with FVII deficiency from 21 unrelated families predominantly of Middle-Eastern extraction. A total of 19 different mutations were identified, of which 12 were novel and 7 had been previously reported. Nine of the 12 novel mutations were missense mutations located in the Gla domain (Ser23Pro), the second epidermal growth factor domain (Cys135Arg) and the catalytic serine protease domain (Arg247Cys, Arg277Cys, Ser282Arg, Pro303Thr, Ser363Ile, Trp364Cys, Trp364Phe), of which five are homozygous. Three novel splice mutations were identified in intron 1a (IVS1a+5), intron 2 (IVS2+1) and intron 6 (IVS6+1). Of the seven previously reported mutations, five were missense mutations of which three are homozygous (Gln100Arg, Arg152Gln, Arg304Gln, Cys310Phe and Thr359Met), one was a 17 bp deletion (10585del17bp) and one was a splice site mutation within intron 7 (IVS7+7). This study has significantly extended the current database of FVII mutations, including the number of known homozygous mutations. Conformational analyses of crystal structures for FVIIa and the FVIIa-tissue factor complex provided likely explanations for the effect of the missense mutations on FVIIa secretion or function. In particular, since 23 missense mutations were located to the serine protease domain, mostly to the region between the catalytic triad and the contact surface with tissue factor, this showed that the orientation of the serine protease domain relative to bound tissue factor in the complex is crucial for functional activity.
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