Determination and control of low-level amino acid misincorporation in human thioredoxin protein produced in a recombinant Escherichia coli production system

Biotechnology and Bioengineering
Robert P HarrisPeter M Kilby

Abstract

Escherichia coli is used extensively in the production of proteins within biotechnology for a number of therapeutic applications. Here, we discuss the production and overexpression of the potential biopharmaceutical human thioredoxin protein (rhTRX) within E. coli. Overexpression of foreign molecules within the cell can put an enormous amount of stress on the translation machinery. This can lead to a misfiring in the construction of a protein resulting in populations differing slightly in amino acid composition. Whilst this may still result in a population of active molecules being expressed, it does present significant problems with molecules that are destined for clinical applications. Amino acid misincorporation of this subset could potentially result in antibodies being raised to these unnatural proteins. Cross-reaction with a patient's endogenous thioredoxin could then lead to an autoimmune phenomena and serious health implications. Generally, the issue of misincorporation appears not to be a routine regulatory concern (see ICH Q6B guidelines). Therefore, amino acid misincorporation may not have been detected, much less explored in the clinic as the occurrence or absence of these random errors is not routinely reported. Us...Continue Reading

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Citations

Jun 13, 2014·Current Opinion in Biotechnology·Robert P Harris, Peter M Kilby
Oct 31, 2012·Biotechnology and Bioengineering·Lauren FeeneyMichael W Laird
Feb 7, 2020·International Journal of Molecular Sciences·Kangsan KimByung-Kwan Cho
Jun 26, 2021·Scientific Reports·Anushikha ThakurRamakrishnan Melarkode Subbaraman

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