Identification of the c.1603A>G and c.1609G>A mutations in the HIF2A gene. (A) Detection of the c.1603A>G and c.1609G>A mutations by PCR-direct sequencing. PCR was performed on total peripheral blood DNA using a set of primers to specifically amplify exon 12 of the HIF2A gene. Sequencing detected a heterozygous A to G change at base 1603 in patient H2 (middle panel) and G to A at base 1609 in patient H5 (bottom panel) as indicated by arrows compared with wild-type sequence (top panel). Shown are nucleotides 1587 to 1615. Bases are as follows: G, black; A, green; T, red; C, blue. (B) Three-dimensional structure of the VHL:ElonginC:ElonginB complex bound to hydroxyproline-564 HIF-1α peptide (residues 556-575). The structure was generated using Cn3D from PDB coordinates (1LM8) deposited by Min et al.18 The positions of hydroxyproline-564 (Hyp-564), Met-568, and Asp-570 are shown (all in bright yellow). The sequences of HIF-2α and HIF-1α at the primary hydroxylation site are compared, with the corresponding HIF-2α residues indicated.
