Engineering a Polyketide Synthase for In Vitro Production of Adipic Acid

Journal Article · Mon Oct 26 04:00:00 EDT 2015 · ACS Synthetic Biology

DOI:https://doi.org/10.1021/acssynbio.5b00153· OSTI ID:1378944

Hagen, Andrew ^[1]; Poust, Sean ^[2]; Rond, Tristan de ^[2]; Fortman, Jeffrey L. ^[1]; Katz, Leonard ^[3]; Petzold, Christopher J. ^[4]; Keasling, Jay D. ^[5]

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1. Univ. of California, Berkeley, CA (United States); U.S. Dept. of Energy, Emeryville, CA (United States)
2. Univ. of California, Berkeley, CA (United States)
3. U.S. Dept. of Energy, Emeryville, CA (United States)
4. U.S. Dept. of Energy, Emeryville, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States)
5. Univ. of California, Berkeley, CA (United States); U.S. Dept. of Energy, Emeryville, CA (United States); Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States)

Polyketides have enormous structural diversity, yet polyketide synthases (PKSs) have thus far been engineered to produce only drug candidates or derivatives thereof. Thousands of other molecules, including commodity and specialty chemicals, could be synthesized using PKSs if composing hybrid PKSs from well-characterized parts derived from natural PKSs was more efficient. Here, using modern mass spectrometry techniques as an essential part of the design–build–test cycle, we engineered a chimeric PKS to enable production one of the most widely used commodity chemicals, adipic acid. To accomplish this, we introduced heterologous reductive domains from various PKS clusters into the borrelidin PKS’ first extension module, which we previously showed produces a 3-hydroxy-adipoyl intermediate when coincubated with the loading module and a succinyl-CoA starter unit. Acyl-ACP intermediate analysis revealed an unexpected bottleneck at the dehydration step, which was overcome by introduction of a carboxyacyl-processing dehydratase domain. Appending a thioesterase to the hybrid PKS enabled the production of free adipic acid. Using acyl-intermediate based techniques to “debug” PKSs as described here, it should one day be possible to engineer chimeric PKSs to produce a variety of existing commodity and specialty chemicals, as well as thousands of chemicals that are difficult to produce from petroleum feedstocks using traditional synthetic chemistry.

• Research Organization: Lawrence Berkeley National Laboratory (LBNL), Berkeley, CA (United States)
• Sponsoring Organization: USDOE Office of Science (SC), Biological and Environmental Research (BER) (SC-23); National Science Foundation (NSF)
• DOE Contract Number: AC02-05CH11231
• OSTI ID: 1378944
• Journal Information: ACS Synthetic Biology, Journal Name: ACS Synthetic Biology Journal Issue: 1 Vol. 5; ISSN 2161-5063
• Publisher: American Chemical Society (ACS)
• Country of Publication: United States
• Language: English

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
60 APPLIED LIFE SCIENCES
adipic acid
polyketide synthase
tandem mass-spectrometry