Expanding Extender Substrate Selection for Unnatural Polyketide Biosynthesis by Acyltransferase Domain Exchange within a Modular Polyketide Synthase
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- Joint BioEnergy Institute, Emeryville, California 94608, United States, Science for Life Laboratory, KTH - Royal Institute of Technology, 17165 Stockholm, Sweden
- Joint BioEnergy Institute, Emeryville, California 94608, United States, Biological Systems and Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, California 94720, United States, Institute of Applied Microbiology, Aachen Biology and Biotechnology, RWTH Aachen University, 52074 Aachen, Germany
- Biological Systems and Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, California 94720, United States, Department of Chemical and Biomolecular Engineering, University of California, Berkeley, California 94720, United States
- Department of Chemical and Biomolecular Engineering, University of California, Berkeley, California 94720, United States, Department of Bioengineering, University of California, Berkeley, California 94720, United States
- Joint BioEnergy Institute, Emeryville, California 94608, United States, Department of Biomaterials and Biomanufacturing, Sandia National Laboratory, Livermore, California 94550, United States
- Joint BioEnergy Institute, Emeryville, California 94608, United States, Biological Systems and Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, California 94720, United States
- Department of Molecular and Cell Biology, University of California, Berkeley, California 94720, United States
- Biological Systems and Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, California 94720, United States, Department of Bioengineering, University of California, Berkeley, California 94720, United States
- Joint BioEnergy Institute, Emeryville, California 94608, United States, Environmental Genomics and Systems Biology Division, Lawrence Berkeley National Laboratory, Berkeley, California 94720, United States
- Joint BioEnergy Institute, Emeryville, California 94608, United States, Molecular Biophysics and Integrated Bioimaging Division, Lawrence Berkeley National Laboratory, Berkeley, California 94720, United States, Department of Bioengineering, University of California, Berkeley, California 94720, United States
- Joint BioEnergy Institute, Emeryville, California 94608, United States, QB3 Institute, University of California, Berkeley, California 94720, United States
- Joint BioEnergy Institute, Emeryville, California 94608, United States, Biological Systems and Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, California 94720, United States, Systems Biology Program, Graduate School of Media and Governance, Keio University, Fujisawa, Kanagawa 252-0882, Japan, Institute of Advanced Biosciences, Keio University, Tsuruoka, Yamagata 997-0017, Japan
- Joint BioEnergy Institute, Emeryville, California 94608, United States, Biological Systems and Engineering Division, Lawrence Berkeley National Laboratory, Berkeley, California 94720, United States, Department of Chemical and Biomolecular Engineering, University of California, Berkeley, California 94720, United States, Department of Bioengineering, University of California, Berkeley, California 94720, United States, QB3 Institute, University of California, Berkeley, California 94720, United States
Modular polyketide synthases (PKSs) are polymerases that employ α-carboxyacyl-CoAs as extender substrates. This enzyme family contains several catalytic modules, where each module is responsible for a single round of polyketide chain extension. Although PKS modules typically use malonyl-CoA or methylmalonyl-CoA for chain elongation, many other malonyl- CoA analogues are used to diversify polyketide structures in nature. Previously, we developed a method to alter an extension substrate of a given module by exchanging an acyltransferase (AT) domain while maintaining protein folding. Here, we report in vitro polyketide biosynthesis by 13 PKSs (the wild-type PKS and 12 AT-exchanged PKSs with unusual ATs) and 14 extender substrates. Our ~200 in vitro reactions resulted in 13 structurally different polyketides, including several polyketides that have not been reported. In some cases, AT-exchanged PKSs produced target polyketides by >100-fold compared to the wild-type PKS. These data also indicate that most unusual AT domains do not incorporate malonyl-CoA and methylmalonyl-CoA but incorporate various rare extender substrates that are equal to in size or slightly larger than natural substrates. We developed a computational workflow to predict the approximate AT substrate range based on active site volumes to support the selection of ATs. These results greatly enhance our understanding of rare AT domains and demonstrate the benefit of using the proposed PKS engineering strategy to produce novel chemicals in vitro.
- Research Organization:
- Univ. of California, Berkeley, CA (United States)
- Sponsoring Organization:
- USDOE Office of Science (SC), Biological and Environmental Research (BER); Formas Mobility
- Grant/Contract Number:
- DEAC02-05CH11231; 2017-00335; AC02-05CH11231
- OSTI ID:
- 1970085
- Alternate ID(s):
- OSTI ID: 1971468
- Journal Information:
- Journal of the American Chemical Society, Journal Name: Journal of the American Chemical Society Vol. 145 Journal Issue: 16; ISSN 0002-7863
- Publisher:
- American Chemical SocietyCopyright Statement
- Country of Publication:
- United States
- Language:
- English
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