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PEGylation of Proteins in Organic Solution: A Case Study for Interferon beta-1b

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National Key Laboratory of Biochemical Engineering, Institute of Process Engineering, Chinese Academy of Sciences, Beijing 100190, China
§ Graduate University of Chinese Academy of Sciences, Beijing 100049, China
*E-mail: [email protected], [email protected]. Tel: +86-10-62561817. Fax: +86-10-62561813.
Cite this: Bioconjugate Chem. 2012, 23, 9, 1812–1820
Publication Date (Web):August 9, 2012
https://doi.org/10.1021/bc300081f
Copyright © 2012 American Chemical Society
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Abstract

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Conventional protein PEGylation is carried out in aqueous solution. However, some hydrophobic proteins seem to be stable in organic solution. In this study, a novel approach of PEGylating IFN-β-1b in an organic solution of 2-butanol (2-BuOH) was investigated. Compared with protein PEGylation in aqueous solution, the overall modification yields increased more than 37%, while the yield of mono-PEGylated products could be increased by 36%. Furthermore, the PEGylated IFN-β-1b, which was obtained in organic solution, demonstrated 18% more antiviral potency than those derived from aqueous solution. The PEGylation step could be directly connected to the previous protein separation step for process integration. Dynamic light scattering (DLS) and atomic force microscope (AFM) analysis revealed that IFN-β-1b formed aggregates both in water and in 2-BuOH solutions. However, the aggregates were much smaller and more homogeneous in 2-BuOH than those in aqueous solution, thereby providing larger solvent accessible protein surfaces, which resulted in a more productive PEGylation process. In addition, the results of circular dichroism (CD), fluorescence spectra, and peptide mapping suggested that the increased bioactivity came from the difference in PEGylation site distribution due to solution environment that induced conformational discrepancy. The results of this study show that PEGylation of IFN-β-1b in organic solution is a facile and efficient process, which might find applications for other hydrophobic proteins.

Cited By


This article is cited by 8 publications.

  1. Shayan Abbasi, Homa Farahani, Hossein Lanjanian, Mohammad Taheri, Loghman Firoozpour, Jamshid Davoodi, Sama Pirkalkhoran, GholamHossein Riazi, Shahriar Pooyan. Site Directed Disulfide PEGylation of Interferon-β-1b with Fork Peptide Linker. Bioconjugate Chemistry 2020, 31 (3) , 708-720. https://doi.org/10.1021/acs.bioconjchem.9b00839
  2. Zhan Zhou, Jing Zhang, Lijing Sun, Guanghui Ma, and Zhiguo Su . Comparison of Site-Specific PEGylations of the N-Terminus of Interferon Beta-1b: Selectivity, Efficiency, and in Vivo/Vitro Activity. Bioconjugate Chemistry 2014, 25 (1) , 138-146. https://doi.org/10.1021/bc400435u
  3. Mohsin Vahid Khan, Syed Mohammad Zakariya, Rizwan Hasan Khan. Protein folding, misfolding and aggregation: A tale of constructive to destructive assembly. International Journal of Biological Macromolecules 2018, 112 , 217-229. https://doi.org/10.1016/j.ijbiomac.2018.01.099
  4. Longfu Xu, Chun Zhang, Qi Wang, Fangxia Guo, Zenglan Li, Yongdong Liu, Zhiguo Su. Oxidized catechol-derived poly (ethylene glycol) for thiol-specific conjugation. Reactive and Functional Polymers 2017, 117 , 97-105. https://doi.org/10.1016/j.reactfunctpolym.2017.06.005
  5. Bahman Khameneh, Mahmoud Reza Jaafari, Mohammad Hassanzadeh-Khayyat, AbdolReza Varasteh, JamshidKhan Chamani, Mehrdad Iranshahi, Hamid Mohammadpanah, Khalil Abnous, Mohammad Reza Saberi. Preparation, characterization and molecular modeling of PEGylated human growth hormone with agonist activity. International Journal of Biological Macromolecules 2015, 80 , 400-409. https://doi.org/10.1016/j.ijbiomac.2015.06.037
  6. Mohsin Vahid Khan, Gulam Rabbani, Mohd Ishtikhar, Shariqua Khan, Gajender Saini, Rizwan Hasan Khan. Non-fluorinated cosolvents: A potent amorphous aggregate inducer of metalloproteinase-conalbumin (ovotransferrin). International Journal of Biological Macromolecules 2015, 78 , 417-428. https://doi.org/10.1016/j.ijbiomac.2015.04.021
  7. Xu-Dong Wang, Hu Teng, Jing-Jing Hu, Zhi-Long Xiu. PEGylation of recombinant hirudin in mixed aqueous–organic solutions. Process Biochemistry 2015, 50 (3) , 367-377. https://doi.org/10.1016/j.procbio.2015.01.007
  8. Fei Peng, Yongdong Liu, Xiunan Li, Lijing Sun, Dawei Zhao, Qingqing Wang, Guanghui Ma, Zhiguo Su. PEGylation of G-CSF in organic solvent markedly increase the efficacy and reactivity through protein unfolding, hydrolysis inhibition and solvent effect. Journal of Biotechnology 2014, 170 , 42-49. https://doi.org/10.1016/j.jbiotec.2013.10.037

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