The mutant forms of GPIIIa containing an alanine instead of a cysteine at amino acid 5 or 435.

(A) DNA sequence analysis of Ala5GPIIIa and Ala435GPIIIa cDNA. The nucleotide sequence was analyzed on both strands using automated sequencing. As indicated, the nucleotide substitutions of TG with GC (GCT) resulted in Cys5Ala and Cys435Ala mutations. (B) Western blot analysis of expressed WT GPIIIa, Ala5GPIIIa, and Ala435GPIIIa. CHO cells (mock) and stable CHO cell lines expressing WT GPIIb-IIIa (WT), GPIIb-Ala5IIIa (Ala5), or GPIIb-Ala435IIIa (Ala435) were solubilized in lysis buffer. Soluble lysates were electrophoresed on 7% SDS-PAGE (left, reduced; right, nonreduced), and subjected to Western blot with rabbit polyclonal antibodies specific for GPIIIa.