A rational design for hepatitis B virus X protein refolding and bioprocess development guided by second virial coefficient studies

Anindya Basu, Wei Ning Chen, Susanna Su Jan Leong

Research output: Contribution to journalArticlepeer-review

7 Citations (Scopus)

Abstract

The hepatitis B virus X (HBx) protein is well known for its role in hepatitis B virus infection that often leads to hepatocellular carcinoma. Despite the clinical importance of HBx, there is little progress in anti-HBx drug development strategies due to shortage of HBx from native sources. Consistent expression of HBx as insoluble inclusion bodies within various expression systems has largely hindered HBx manufacturing via economical biosynthesis routes. Confronted by this roadblock, this study aims to quantitatively understand HBx protein behaviour in solution that will guide the rational development of a refolding-based bioprocess for HBx production. Second virial coefficient (SVC) measurements were employed to study the effects of varying physicochemical parameters on HBx intermolecular protein interaction. The SVC results suggest that covalent HBx aggregates play a key role in protein destabilisation during refolding. The use of an SVC-optimised refolding environment yielded bioactive and soluble HBx proteins from the denatured-reduced inclusion body state. This study provides new knowledge on HBx solubility behaviour in vitro, which is important in structure-function elucidation behaviour of this hydrophobic protein. Importantly, a rational refolding-based Escherichia coli bioprocess that can deliver purified and soluble HBx at large scale is successfully developed, which opens the way for rapid preparation of soluble HBx for further clinical and characterisation studies.

Original languageEnglish
Pages (from-to)181-191
Number of pages11
JournalApplied Microbiology and Biotechnology
Volume90
Issue number1
DOIs
Publication statusPublished - Apr 2011
Externally publishedYes

ASJC Scopus Subject Areas

  • Biotechnology
  • Applied Microbiology and Biotechnology

Keywords

  • Bioprocess
  • Hepatitis B virus X protein
  • Refolding
  • Second virial coefficient
  • Solubility

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