Cell-Free Protein Synthesis for High-Throughput Biosynthetic Pathway Prototyping

Rasor, Blake J.; Vögeli, Bastian; Jewett, Michael C.; Karim, Ashty S.

doi:10.1007/978-1-0716-1998-8_12

Title: Cell-Free Protein Synthesis for High-Throughput Biosynthetic Pathway Prototyping

Journal Article · 04 January 2022 · Methods in Molecular Biology

DOI: https://doi.org/10.1007/978-1-0716-1998-8_12 · OSTI ID:2298963

Rasor, Blake J. ^[1]; Vögeli, Bastian ^[2]; Jewett, Michael C. ^[2]; Karim, Ashty S. ^[2]

Northwestern University, Evanston, IL (United States); Northwestern University
Northwestern University, Evanston, IL (United States)

Biological systems provide a sustainable and complimentary approach to synthesizing useful chemical products. Metabolic engineers seeking to establish economically viable biosynthesis platforms strive to increase product titers, rates, and yields. Despite continued advances in genetic tools and metabolic engineering techniques, cellular workflows remain limited in throughput. It may take months to test dozens of unique pathway designs even in a robust model organism, such as Escherichia coli. In contrast, cell-free protein synthesis enables the rapid generation of enzyme libraries that can be combined to reconstitute metabolic pathways in vitro for biochemical synthesis in days rather than weeks. Cell-free reactions thereby enable comparison of hundreds to thousands of unique combinations of enzyme homologs and concentrations, which can quickly identify the most productive pathway variants to test in vivo or further characterize in vitro. Furthermore, this cell-free pathway prototyping strategy provides a complementary approach to accelerate cellular metabolic engineering efforts toward highly productive strains for metabolite production.

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Research Organization:: Northwestern University, Evanston, IL (United States)

Sponsoring Organization:: USDOE

Grant/Contract Number:: SC0018249

OSTI ID:: 2298963

Journal Information:: Methods in Molecular Biology, Journal Name: Methods in Molecular Biology Vol. 2433; ISSN 1064-3745

Publisher:: SpringerCopyright Statement

Country of Publication:: United States

Language:: English

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Title: Cell-Free Protein Synthesis for High-Throughput Biosynthetic Pathway Prototyping

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