Strategies for optimization of heterologous protein expression in E. coli: Roadblocks and reinforcements

Kaur, Jagdeep; Kumar, Arbind

doi:10.1016/j.ijbiomac.2017.08.080

Cited by 314 publications

(232 citation statements)

References 277 publications

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“…However, one obstacle to the expression of recombinant protein in E. coli is inclusion body formation, which can reach 70% of the total recombinant protein (Kane and Hartley 1988;Zhong et al 2011), leading to relatively low levels of active heterologous protein. To overcome the high concentration of inclusion bodies, the influence of genetic approaches and induction conditions on the yield of active recombinant protein has been recognized by researchers and industrial communities (Hockney 1994;Kaur et al 2018). Genetic approaches such as fusion protein expression (Malhotra 2009) and co-expression of molecular chaperones (Chou 2007) were used to enhance soluble heterologous protein expression.…”

Section: Introductionmentioning

confidence: 99%

High-level expression of Aerococcus viridans pyruvate oxidase in Escherichia coli by optimization of vectors and induction conditions

Zhao

Zhang

2018

Lett Appl Microbiol

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This study demonstrates that a highly soluble pyruvate oxidase can be obtained in recombinant Escherichia coli by optimizing vectors and induction conditions. The pyruvate oxidase yield achieved is the highest reported so far, which provides a convenient and cost-saving way to produce pyruvate oxidase. This research promotes pyruvate oxidase application in the pharmaceutical and biochemical industries.

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Section: Introductionmentioning

confidence: 99%

High-level expression of Aerococcus viridans pyruvate oxidase in Escherichia coli by optimization of vectors and induction conditions

Zhao

Zhang

2018

Lett Appl Microbiol

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“…Although E.coli expression system has been used widely for recombinant proteins production in laboratory and industrial scale due to its simplicity and economy, however, the solubility of target proteins is low [24][25][26] .…”

Section: Discussionmentioning

confidence: 99%

Enhanced production of Canine parvovirus VP2 protein using improved Baculovirus expression system and its immunogenicity

Chang¹,

Liu²,

Chen³

et al. 2020

Preprint

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Background : Canine parvovirus (CPV) is now recognized as a serious threat to dog industry worldwide. Vaccination remains the principal tool to control CPV infection. However, due to low yield, production of VP2 protein of CPV in baculovirus expression system remains challenging. The aim of this study was to increase the VP2 protein production by using a improved baculovirus expression system (Multibac) and evaluate the immunogenicity of the purified VP2 protein in mice. Results: The results showed that CPV VP2 protein was successfully expressed in the improved baculovirus expression system efficiently. A high level of expression of the full length VP2 protein was achieved using our modified system. The recombinant virus carrying two copies of VP2 showed the highest expression level, with a productivity of 186 mg/L, which is about 1.4-1.6 fold that of the recombinant viruses carrying only one copy. The purified protein could react with Mouse anti-His tag monoclonal antibody and Rabbit anti-VP2 polyclonal antibody with good reactogenicity. The mice were then immunized with purified full length VP2 protein to evaluate its immunogenicity. After vaccination, VP2 protein could induce the mice produce high level of hemagglutination inhibition antibodies. Conclusions: Full length CPV VP2 protein was successfully expressed at high level and purified efficiently. And it stimulated mice to produce high level of antibody. The full length VP2 expressed in this study could be used as a putative economic and efficient subunit vaccine against CPV infection.

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“…In the past, many protocols aimed at avoiding IB formation during heterologous protein production. These so-called midstream approaches, like chaperone co-expression, reduced growth temperature, expression tuning, strains that promote soluble POI production, cofactor addition to the medium, fusion proteins and translocation to the periplasm, are extensively discussed elsewhere (Basu et al 2011; Burgess 2009; García-Fruitós et al 2012; Kaur et al 2017; Sørensen and Mortensen 2005). Albeit, these attempts rarely resulted in sufficient yields of soluble POI to be relevant for industrial purposes.…”

Section: What Are Inclusion Bodies?mentioning

confidence: 99%

“…Finally, the refolded POI might undergo a purification step to further increase purity. A more detailed IB process description can be found elsewhere (Basu et al 2011; Burgess 2009; Eggenreich et al 2016; Hoffmann et al 2017; Kaur et al 2017; Rathore et al 2013; Singh et al 2015). …”

Section: What Are Inclusion Bodies?mentioning

confidence: 99%

Wanted: more monitoring and control during inclusion body processing

Humer

Spadiut

2018

World J Microbiol Biotechnol

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Inclusion bodies (IBs) are insoluble aggregates of misfolded protein in Escherichia coli. Against the outdated belief that the production of IBs should be avoided during recombinant protein production, quite a number of recombinant products are currently produced as IBs, which are then processed to give correctly folded and soluble product. However, this processing is quite cumbersome comprising IB wash, IB solubilization and refolding. To date, IB processing often happens rather uncontrolled and relies on empiricism rather than sound process understanding. In this mini review we describe current efforts to introduce more monitoring and control in IB processes, focusing on the refolding step, and thus generate process understanding and knowledge.

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Strategies for optimization of heterologous protein expression in E. coli: Roadblocks and reinforcements

Cited by 314 publications

References 277 publications

High-level expression of Aerococcus viridans pyruvate oxidase in Escherichia coli by optimization of vectors and induction conditions

High-level expression of Aerococcus viridans pyruvate oxidase in Escherichia coli by optimization of vectors and induction conditions

Enhanced production of Canine parvovirus VP2 protein using improved Baculovirus expression system and its immunogenicity

Wanted: more monitoring and control during inclusion body processing

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