GPIX_E convex loops participate in direct interaction with GPIbβ_E in the full-length complex. (A) SDS gels showing the lack of disruptive effects of the L82A or D86E mutation on expression, secretion, and folding of HA-GPIbβ_Eabc. Residue numbers of both mutations are in the context of GPIX. The other annotations follow those of Figure 2C. (B) Relative surface expression levels of HA-GPIbβ_Eabc-GPIX_TC, either wild-type or containing the indicated mutation, in the absence (gray column) or presence (white column) of coexpressing GPIbβ in transfected CHO cells. HA-GPIbβ_Eabc-GPIX_TC is a protein that contains N-terminally HA-tagged GPIbβ_Eabc and GPIX TM and cytoplasmic domains.¹⁶  The expression level was measured by flow cytometry with the use of anti-HA mAb and quantified as mean fluorescence intensity, which was normalized with that in cells transfected with wild-type GPIb-IX (GPIbα/GPIbβ/HA-GPIX) being 100% and those in cells transfected with sham vectors 0%.¹⁶  Note that the enhancement of HA-GPIbβ_Eabc-GPIX_TC surface expression by GPIbβ is significantly diminished by both mutations. The data are presented as mean ± SD (n = 4). *P < .01. (C) Relative surface expression levels of GPIX, either wild-type or containing the indicated mutation, in the absence (gray column) or presence (white column) of coexpressing GPIbβ in transfected CHO cells. GPIX surface expression level was measured by flow cytometry with the use of anti-GPIX mAb FMC25 and analyzed as described above (n = 3).