An important risk factor for
thrombosis is the polymorphism R506Q in
factor V that causes resistance of
factor Va to proteolytic inactivation by activated
protein C (APC). To study the potential influence of the
carbohydrate moieties of
factor Va on its inactivation by APC,
factor V was subjected to mild deglycosylation (
neuraminidase plus
N-glycanase) under nondenaturing conditions. The
APC resistance ratio values (ratio of activated partial thromboplastin time [APTT] clotting times with and without APC) of the treated
factor V were increased (2.4 to 3.4) as measured in APTT assays.
O-glycanase treatment of
factor V did not change the
APC resistance ratio. The procoagulant activity of
factor V as well as its activation by
thrombin was not affected by mild deglycosylation. Treatment of
factor V with
neuraminidase and
N-glycanase mainly altered the electrophoretic mobility of the
factor Va heavy chain, whereas treatment with
O-glycanase changed the mobility of the connecting region. This suggests that the removal of the N-linked
carbohydrates from the heavy chain of
factor Va, which is the substrate for APC, is responsible for the increase in susceptibility to inactivation by APC. Thus, variability in
carbohydrate could account for some of the known variability in
APC resistance ratios, including the presence of borderline or low
APC resistance ratios among patients who lack the R506Q mutation.