Factor VII Deficiency Caused by a Structural Variant N57D of the
First Epidermal Growth Factor Domain
Blair J.N. Leonard,
Qi Chen,
Morris A. Blajchman,
Frederick A. Ofosu,
Sampath Sridhara,
Daniel Yang, and
Bryan J. Clarke
From the Departments of Pathology, Biochemistry, and Biomedical
Sciences, McMaster University; and the Canadian Red Cross Society,
Hamilton, Ontario, Canada.
We have previously described a kindred with factor VII (FVII)
deficiency whose members exhibited reduced procoagulant activity relative to FVII antigen concentration. In this report,
the molecular genetic basis of the FVII defect has been determined to
be a heterozygous substitution of Asp for Asn at position 57 in the
first epidermal growth factor (EGF) domain. Recombinant FVII (N57D)
cDNA was created by site-directed mutagenesis and transiently expressed
in human 293 cells. The transfected cells synthesized an
immunoprecipitable protein with an apparent molecular weight of 50 kD. Quantitation of expression by FVII enzyme-linked
immunosorbent assay indicated that mutant protein yields were
consistently low, typically 10% to 30% of wild-type FVII. FVII (N57D)
protein did not accumulate intracellularly, and Northern blot analysis
indicated equivalent FVII mRNA levels in 293 cells expressing either
wild-type FVII or FVII (N57D). Secreted FVII (N57D) protein did not
bind tissue factor, exhibited no procoagulant activity, and failed to
bind a conformation-dependent monoclonal antibody specific for the first EGF domain of FVII. Molecular modeling of the first EGF domain of
FVII predicted that the N57D amino acid substitution would disrupt
tertiary bonding structure. We conclude that the N57D mutation affects
folding of the first EGF domain of FVII resulting in decreased cellular
secretion of a mutant FVII molecule, which is unable to bind tissue
factor and is therefore biologically inactive.
Blood, Vol. 91 No. 1 (January 1), 1998:
pp. 142-148
© 1998 by The American Society of Hematology.
