Effect of GGCX’s C terminus on BGP binding. (A) Interaction of the R704X mutant with 3 VKD reporter proteins in live cells probed by DSS cross-linking. The R704X mutant and MBP fusions of the VKD reporter proteins (FIXgla, MGP, BGP) were transiently coexpressed in GGCX-deficient HEK293 cells for DSS cross-linking, as described in Figure 4B. The cross-linked bands of GGCX and its substrate were marked with asterisks. (B) Interaction of the R704X mutant with 3 VKD reporter proteins in live cells probed by the Venus-based BiFC assay. R704X-VN and the reporter protein fused with VC were transiently coexpressed in GGCX-deficient HEK293 cells. Forty-eight hours later, the fluorescence signal was determined directly from the transfected live cells. Data are presented as the mean ± SD of 3 independent experiments (n = 3). *P < .001. (C) Interaction of GGCX C-terminal truncation mutations with BGP probed by DSS cross-linking, as described in panel A. (D) BGP carboxylation by GGCX C-terminal truncation mutations in GGCX-deficient HEK293 cells, as described in Figure 2B. Data are presented as the mean ± SD of 3 independent experiments (n = 3). *P < .001.