Gene Construction, Expression, Purification and Activity Analysis of Anti-platelet Aggregation Peptide RGD226
YANG Xing-wen 1), LIU Jian 2), TANG Jian-guo 1)*
( 1)National Laboratory of Protein Engineering and Plant Gene Engineering, College of Life Sciences, Peking University, Beijing 100871,China; 2)Institute of Medicine Plant Development,
Abstract：Human proinsulin mutant ([A6-Ser, A11-Ser] proinsulin) gene between B28 and A1 sites was replaced by a gene encoding a peptide of 13 amino acids, SRVARGDWNDDYS, by PCR method. The peptide with an RGD (Arg-Gly-Asp) motif was originated from disintegrin eriststatin. The constructed RGD226 gene was overexpressed in E. coli. RGD226 peptide was purified by Sephadex G-50 chromatography and by Resource TM reverse phase chromatography successively. The RGD226 peptide inhibits ADP-induced human platelet aggregation with an IC 50 of 22.3 μmol/L and keeps 15.1% of immunity of [A6-Ser, A11-Ser] proinsulin, but with 0.1% of receptor binding activity of [A6-Ser, A11-Ser] proinsulin. The in vivo results suggest that the RGD226 peptide can obviously delay the time of bleeding in mice.
