A novel engineered interchain disulfide bond in the constant region enhances the thermostability of adalimumab Fab

Hitomi Nakamura, Naoko Oda-Ueda, Tadashi Ueda, Takatoshi Ohkuri

研究成果: ジャーナルへの寄稿記事

3 引用 (Scopus)

抄録

We constructed a system for expressing the Fab of the therapeutic human monoclonal antibody adalimumab at a yield of 20 mg/L in the methylotrophic yeast Pichia pastoris. To examine the contribution of interchain disulfide bonds to conformational stability, we prepared adalimumab Fab from which the interchain disulfide bond at the C-terminal region at both the CH1 and CL domains was deleted by substitution of Cys with Ala (FabΔSS). DSC measurements showed that the Tm values of FabΔSS were approximately 5 °C lower than those of wild-type Fab, suggesting that the interchain disulfide bond contributes to conformational thermostability. Using computer simulations, we designed a novel interchain disulfide bond outside the C-terminal region to increase the stability of FabΔSS. The resulting Fab (mutSS FabΔSS) had the mutations H:V177C and L:Q160C in FabΔSS, confirming the formation of the disulfide bond between CH1 and CL. The thermostability of mutSS FabΔSS was approximately 5 °C higher than that of FabΔSS. Therefore, the introduction of the designed interchain disulfide bond enhanced the thermostability of FabΔSS and mitigated the destabilization caused by partial reduction of the interchain disulfide bond at the C-terminal region, which occurs in site-specific modification such as PEGylation.

元の言語英語
ページ(範囲)7-11
ページ数5
ジャーナルBiochemical and Biophysical Research Communications
495
発行部数1
DOI
出版物ステータス出版済み - 1 1 2018

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Disulfides
Pichia
Adalimumab
Computer Simulation
Yeast
Substitution reactions
Yeasts
Monoclonal Antibodies
Mutation
Computer simulation

All Science Journal Classification (ASJC) codes

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

これを引用

A novel engineered interchain disulfide bond in the constant region enhances the thermostability of adalimumab Fab. / Nakamura, Hitomi; Oda-Ueda, Naoko; Ueda, Tadashi; Ohkuri, Takatoshi.

:: Biochemical and Biophysical Research Communications, 巻 495, 番号 1, 01.01.2018, p. 7-11.

研究成果: ジャーナルへの寄稿記事

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