WATER-SOLUBLE RECOMBINANT N-MYC PROTEIN EXPRESSED IN ESCHERICHIA-COLI

Background - The importance of determining N-Myc oncoprotein rather th an genomic N-myc amplification in neuroblastoma has been emphasized be cause of the discrepancies between N-myc amplification and either its expression in some neuroblastoma cell lines or prognosis in some stage IV patients, In order to quantify the expression of N-Myc oncoprotein , a standard protein is required, Methods - An expression plasmid carr ying the coding region of the N-myc gene was constructed and induced b y isopropyl-beta-D-thiogalactopyranoside (IPTG) tea express recombinan t human N-Myc protein (rNMyc, codons 19 to 464), rNMyc was purified by nickel-resin affinity chromatography, The properties of expressed rNM yc were analyzed by sodium dodecylsulfate-10% polyacrylamide gel elect rophoresis with Coomassie brilliant blue staining and immunoblotting w ith anti-N-Myc protein antibodies recognizing different epitopes, Resu lts - Optimal expression of rNMyc was obtained at Ih after IPTG induct ion, rNMyc was water-soluble, had a molecular weight of 38 kDa and rea cted with anti-N-Myc antibodies. Conclusion - Newly obtained rNMyc is water-soluble and covers almost the full length of N-Myc oncoprotein.