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Tools to cope with difficult-to-express proteins

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Abstract

The identification of DNA coding sequences contained in the genome of many organisms coupled to the use of high throughput approaches has fueled the field of recombinant protein production. Apart from basic research interests, the growing relevance of this field is highlighted by the global sales of the top ten biopharmaceuticals on the market, which exceeds the trillion USD in a steady increasing tendency. Therefore, the demand of biological compounds seems to have a long run on the market. One of the most popular expression systems is based on Escherichia coli cells which apart from being cost-effective counts with a large selection of resources. However, a significant percentage of the genes of interest are not efficiently expressed in this system, or the expressed proteins are accumulated within aggregates, degraded or lacking the desired biological activity, being finally discarded. In some instances, expressing the gene in a homologous expression system might alleviate those drawbacks but then the process usually increases in complexity and is not as cost-effective as the prokaryotic systems. An increasing toolbox is available to approach the production and purification of those difficult-to-express proteins, including different expression systems, promoters with different strengths, cultivation media and conditions, solubilization tags and chaperone coexpression, among others. However, in most cases, the process follows a non-integrative trial and error strategy with discrete success. This review is focused on the design of the whole process by using an integrative approach, taken into account the accumulated knowledge of the pivotal factors that affect any of the key processes, in an attempt to rationalize the efforts made in this appealing field.

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Acknowledgments

The authors acknowledge the financial support received for funding our research on protein-based nanotherapeutics and biomaterials from Agència de Gestió d’Ajuts Universitaris i de Recerca (AGAUR, 2014SGR-132), FIS (PS09-00165 and PI12/00327), Fundació La Marató de TV3 (TV32009-101235, TV32013-132031, and TV32013-133930), INIA (RTA2012-00028-C02-02), MINECO (BFU2010-17450, ACI2009-0919, IT2009-0021, BIO2013-41019-P, LIPOCELL TT02, and TERARMET RTC-2014-2207-1), and from the Centro de Investigación Biomédica en Red (CIBER) de Bioingeniería, Biomateriales y Nanomedicina (NANOLYSO, NANOPROTHER, and PENTRI projects). CIBER de Bioingeniería, Biomateriales y Nanomedicina (CIBER-BBN) is an initiative funded by the VI National R&D&i Plan 2008–2011, Iniciativa Ingenio 2010, Consolider Program, CIBER Actions, and financed by the Instituto de Salud Carlos III with assistance from the European Regional Development Fund.

We also acknowledge the ICTS “NANBIOSIS”, more specifically to the Protein Production Platform of CIBER in Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN)/IBB, at the UAB SepBioES scientific-technical service (http://www.nanbiosis.es/unit/u1-protein-production-platform-ppp/).

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  1. Institut de Biotecnologia i de Biomedicina, Universitat Autònoma de Barcelona, Bellaterra, 08193, Barcelona, Spain

    Paolo Saccardo, José Luís Corchero & Neus Ferrer-Miralles

  2. CIBER de Bioingeniería, Biomateriales y Nanomedicina (CIBER-BBN), Bellaterra, 08193, Barcelona, Spain

    Paolo Saccardo, José Luís Corchero & Neus Ferrer-Miralles

  3. Departament de Genètica i de Microbiologia, Universitat Autònoma de Barcelona, Bellaterra, 08193, Barcelona, Spain

    José Luís Corchero & Neus Ferrer-Miralles

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  1. Paolo Saccardo
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Correspondence to Neus Ferrer-Miralles.

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Saccardo, P., Corchero, J.L. & Ferrer-Miralles, N. Tools to cope with difficult-to-express proteins. Appl Microbiol Biotechnol 100, 4347–4355 (2016). https://doi.org/10.1007/s00253-016-7514-8

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