Absence of Vitamin K-Dependent γ-Carboxylation in Human Periostin Extracted from Fibrotic Lung or Secreted from a Cell Line Engineered to Optimize γ-Carboxylation
Fig 5
Recombinant Vitamin K-dependent coagulation factor VII produced in HEK293VKOR cells is γ-carboxylated whereas PN is not.
(A) Conditioned media containing 10 μg/mL vitamin K from HEK293VKOR un-transfected (293) or transfected with PN0 (PN0), 30 μl, or 0.5 pmol of purified factor VII (fVII) were examined in Western blot. Mouse anti-PN (left) reacted strongly with media of cells transfected with PNO but not with control media. Both media were negative when probed with the mouse anti-Gla (right), but purified fVII reacted strongly. (B) Western blot analysis of purified fVII and PN0 expressed by HEK293VKOR cells in the presence of vitamin K. Varying amounts, 10.0, 1.0 and 0.1 pmol, of recombinant protein were probed with anti-PN (left) and anti-Gla (right). The anti-Gla antibody reacted with all inputs tested of fVII but not with PN.