Figure 8.

Site mutagenesis and structural modeling analysis further indicated that N-glycans might directly modulate Klotho's protein folding and co-receptor function. A, a single N-linked site mutant of sKlotho was transiently transfected into HEK293 cells as described under “Experimental procedures.” Both conditioned medium (CM) and cell pellets were analyzed by SDS-PAGE and immunoblotted with anti-His₄ antibody (Qiagen, Germantown, MD). B, structural modeling of sKlotho/FGFR1c/FGF23 complex. Structure from Protein Data Bank entry 5W21 (32) was modeled with extended sugar chains from Fc glycan in Protein Data Bank entry 5VGP.