Premium
An Approach for Increasing the Mass Recovery of Proteins Derived from Inclusion Bodies in Biotechnology
Author(s) -
Wu Dan,
Wang Chaozhan,
Geng Xindu
Publication year - 2007
Publication title -
biotechnology progress
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.572
H-Index - 129
eISSN - 1520-6033
pISSN - 8756-7938
DOI - 10.1021/bp060257m
Subject(s) - chromatography , chemistry , peg ratio , hydrophilic interaction chromatography , molecular mass , high performance liquid chromatography , biochemistry , enzyme , finance , economics
A method for increasing the mass recovery of therapeutic proteins produced by E. coli using liquid chromatography was investigated. Recombinant human interferon‐γ (rhIFN‐γ) produced by E. coli was selected as a model therapeutic protein, and hydrophobic interaction chromatography (HIC) was performed as a model for liquid chromatography. Using seven types of stationary phase hydrophobic interaction chromatography (STHIC) with different end groups, the effect of the stationary phase on the mass recovery during protein folding by liquid chromatography (LC) and the causes of mass loss of rhIFN‐γ during its folding with simultaneous purification were investigated. Also strategies for increasing mass recovery are proposed. The results demonstrate that the mass recovery of rhIFN‐γ increases with the decreasing hydrophobicity for six STHIC with end groups of PEG‐200, PEG‐400, PEG‐600, PEG‐1000, furfural, and phenyl, except for PEG‐1000. However, for oxethyl and PEG‐600, even though the same diol end group is bonded to PEG‐600, so long as the PEG‐600 is modified by acetyl chloride, it can effectively enhance the mass and bioactivity recovery of rhIFN‐γ compared to the PEG‐600 column. The effect of sample size including both mass and volume on the mass recovery of the rhIFN‐γ was also investigated. Last, redissolving the target protein that has irreversibly adsorbed to the stationary phase and re‐injecting it onto the column is an approach for increasing mass recovery.