Journal of virological methods, 2014, 01, 55─62

Alternative purification method for recombinant measles viral nucleoprotein expresses in insect cells by ion-exchange chromatography

HanSaem Lee, Y Kim, J Yang, H Yoon, S Kim, K Kim

Abstract

Recombinant measles virus nucleoproteins (rMeV N) and fusion (F) proteins were characterized as major antigenic proteins expressed in insect cells mediated by recombinant baculoviruses (rBVs). Band intensities was analyzed by Western blotting to recognize IgG and IgM antibodies against the rMeV N and F proteins in human sera and cerebrospinal fluids (CSFs) from patients with mealses infections. Positive results from the blots using the rMeV N were consistent with the results of enzyme-linked immunosorbent assays (ELISAs) in which whole viral proteins were used as antigens. Human sera and CSFs reacted more strongly with the rMeV N than rMeV F proteins prepared in an identical expression system. For efficient and reliable purification, ion-exchange chromatography using Source Q anion resin was applied, and high-purity rMeV N protein was harvested. To characterize the similarity with the native viral protein to purified N protein, structural mimicry of purified recombinant proteins with intact rMeV N was shown through transmission electron microscopy, and the truncation and the phosphorylation status of the expressed protein were analyzed. These results suggest that the rMeV N purified by ion-exchange chromatography has features similar to those of na？ve N including a self-assembled structure, phosphorylation and antigenic function. Thus, these expression and purification methods can be applied to the large-scale production of rMeV N, which is essential for the development of new diagnostic tools and vaccines for acute and chronic MeV infections.

• ISBN or ISSN: 0166-0934

• 본 연구는 질병관리본부 연구개발과제(과제번호 2010-감염-2) 연구비를 지원받아 수행되었습니다.
• This research was supported by a fund(code 2010-Infectious diseases-2) by Research of Korea Centers for Disease Control and Prevention.