RESUMO
OBJECTIVE: To explore the effect of glycoprotein (GP) alphaIIb Ala477Pro(A446P) mutation on the expression of integrin alphaIIbbeta3. METHODS: The alphaIIbA477P (A446P) eukaryotic expression plasmid alphaIIbA477P (A446P)-pcDNA3.1(+) was constructed by site-directed mutagenesis. Cos-7 cells were transfected with the mutational plasmid or the normal plasmid alphaIIb-pcDNA3.1(+) with beta3-pcDNA3.1(+). The expression of alphaIIbA477P (A446P)was tested with the RT-PCR, Western blot, and flow cytometry. RESULTS: RT-PCR revealed the transcriptional script of alphaIIbA477P (A446P). Western blot showed the expression of alphaIIbA477P (A446P) protein. Flow cytometry demonstrated that the expression of alphaIIbA477P (A446P)beta3 on the membrane was only 12.95% of the normal alphaIIbbeta3 complex. CONCLUSION: alphaIIbA477P (A446P) mutation distinctly reduces the expression of alphaIIbbeta3 complex on the membrane. This mutation may interfere the formation of alphaIIbbeta3 complex or impair the proper conformation of alphaIIb subunit.
