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Affibody Scaffolds Improve Sesquiterpene Production in Saccharomyces cerevisiae
KTH, School of Biotechnology (BIO), Proteomics and Nanobiotechnology. KTH, Centres, Science for Life Laboratory, SciLifeLab.ORCID iD: 0000-0002-2430-2682
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2017 (English)In: ACS Photonics, ISSN 2186-2311, E-ISSN 2161-5063, Vol. 6, no 1, 19-28 p.Article in journal (Refereed) Published
Abstract [en]

Enzyme fusions have been widely used as a tool in metabolic engineering to increase pathway efficiency by reducing substrate loss and accumulation of toxic intermediates. Alternatively, enzymes can be colocalized through attachment to a synthetic scaffold via noncovalent interactions. Here we describe the use of affibodies for enzyme tagging and scaffolding. The scaffolding is based on the recognition of affibodies to their anti-idiotypic partners in vivo, and was first employed for colocalization of farnesyl diphosphate synthase and farnesene synthase in S. cerevisiae. Different parameters were modulated to improve the system, and the enzyme:scaffold ratio was most critical for its functionality. Ultimately, the yield of farnesene on glucose YSFar could be improved by 135% in fed-batch cultivations using a 2-site affibody scaffold. The scaffolding strategy was then extended to a three-enzyme polyhydroxybutyrate (PHB) pathway, heterologously expressed in E. coli. Within a narrow range of enzyme and scaffold induction, the affibody tagging and scaffolding increased PHB production 7-fold. This work demonstrates how the versatile affibody can be used for metabolic engineering purposes.

Place, publisher, year, edition, pages
American Chemical Society (ACS), 2017. Vol. 6, no 1, 19-28 p.
Keyword [en]
affibodies, biofuels, isoprenoids, metabolic engineering, PHB, yeast
National Category
Biochemistry and Molecular Biology
Identifiers
URN: urn:nbn:se:kth:diva-208453DOI: 10.1021/acssynbio.6b00109ISI: 000392575700003PubMedID: 27560952ScopusID: 2-s2.0-85013082953OAI: oai:DiVA.org:kth-208453DiVA: diva2:1106330
Funder
Science for Life Laboratory - a national resource center for high-throughput molecular bioscienceKnut and Alice Wallenberg FoundationSwedish Research Council FormasNovo Nordisk
Note

QC 20170607

Available from: 2017-06-07 Created: 2017-06-07 Last updated: 2017-06-07Bibliographically approved

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