Improving the Productivity of Single-Chain Fv Antibody Against c-Met by Rearranging the Order of its Variable Domains

  • Kim, Yu-Jin (Department of Polymer Science and Chemical Engineering, Pusan National University) ;
  • Neelamegam, Rameshkumar (Department of Polymer Science and Chemical Engineering, Pusan National University) ;
  • Heo, Mi-Ae (Department of Polymer Science and Chemical Engineering, Pusan National University) ;
  • Edwardraja, Selvakumar (Department of Polymer Science and Chemical Engineering, Pusan National University) ;
  • Paik, Hyun-Jong (Department of Polymer Science and Chemical Engineering, Pusan National University) ;
  • Lee, Sun-Gu (Department of Polymer Science and Chemical Engineering, Pusan National University)
  • Published : 2008.06.30

Abstract

Single-chain Fv (scFv) antibody against c-Met is expected to be employed in clinical treatment or imaging of cancer cells owing to the important biological roles of c-Met in the proliferation of malignancies. Here, we show that the productivity of scFv against c-Met in Escherichia coli is significantly influenced by the orientation of its variable domains. We generated anti-c-Met scFv antibodies with two different domain orders (i.e., $V_L$-linker-$V_H$ and $V_H$-linker-$V_L$), expressed them in the cytoplasm of E. coli trx/gor deleted mutant, and compared their specific activities as well as their productivities. Productivity of total and functional anti-c-Met scFv with $V_H/V_L$ orientation was more than five times higher than that with $V_L/V_H$ format. Coexpression of DsbC enhanced the yield of soluble amounts of anti-c-Met scFv protein for both constructs. The purified scFv antibodies of the two different formats exhibited almost the same antigen-binding activities. We also compared the productivities and specific activities of anti-c-Met diabodies with $V_H/V_L$ or $V_L/V_H$ formats and obtained similar results to the case of scFv antibodies.

Keywords

References

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