Merger of Whole Cell Biocatalysis with Organocatalysis Upgrades Alcohol Feedstocks in a Mild, Aqueous, One-Pot Process

Significance 

Chemical synthesis links a series of chemical reactions together to yield a target product. This is the most common way to produce high value industrial chemicals. An emerging strategy to improve the sustainability of chemical production is to employ biosynthetic methods. Biosynthetic methods have the potential to deliver value-added chemicals from renewable feedstocks at mild temperatures, in aqueous solvent conditions, and in a single pot. Because of these features, biosynthetic processes minimize energy usage and waste streams and, ideally, obviate the need to isolate chemical intermediates. However, despite major advances in metabolic engineering and synthetic biology, the rapid engineering of microbes to deliver high yields and titers of target compounds remains as a challenge. For instance, alteration of one metabolic pathway may have unexpected ramifications in other pathways or the metabolic intermediates and end products build up and become toxic to organisms or trigger feedback inhibition. In-situ product recovery, which is used to extract target products, is appealing but requires that the desired chemical products effectively partition into biocompatible extractants.

To address the aforementioned system challenges, a team of researchers from the Department of Chemistry at Colorado School of Mines, including Ph.D. candidate Kelsey Stewart and Ms. Emily Hicks and led by Professor Dylan Domaille developed a new strategy for biosynthesis. Rather than using overexpressed enzymes to drive flux to a target product, they hypothesized that biocompatible chemical catalysts could play the role of overexpressed enzymes to intercept metabolic intermediates and redirect flux toward target chemicals. Their work is currently published in the research journal, ACS Sustainable Chemistry & Engineering.

In their reported system, a Gram negative non-pathogenic bacteria, Gluconobacter oxidans, was used to oxidize aliphatic alcohols to aliphatic aldehydes. When lysine, a biocompatible aldol catalyst, was added to the culture media, the aldehyde was dimerized to its aldol condensation product. A feedstock substrate of n-butanol delivered good yields of the high-volume chemical, 2-ethyl-2-hexenal (2-EH), in aqueous buffer at mild temperature. This is in sharp contrast to the existing method of 2-EH production, which involves high temperatures and creates large volumes of basic waste streams. A biocompatible isooctane extractant selectively removed the C2-EH product as it formed, driving flux toward the target product.

Product analysis of aqueous and organic phases revealed that in the absence of catalyst, only n-butanoic acid, which is the product of continued microbial oxidation, was produced. This analysis revealed that the in-situ lysine catalyst redirected metabolic flux to 2-EH, minimizing production of the undesirable n-butanoic acid. Moreover, the authors were able to show that their approach upgraded a range of C2−C6 n-alkyl alcohols in both self- and crossed aldol-type reactions.

In summary, the study presented a mild, aqueous synthesis approach of the high-volume chemical 2-EH from n-butanol. Further, it also demonstrated that the presented methodology was applicable to the self- and crossed aldol type couplings of C2−C6 substrates. In a statement to Advances in Engineering, Professor Dylan W. Domaille pointed out that their work revealed an exciting new strategy of flux redirection that can be used to expand the scope of products from biosynthetic processes.

Merger of Whole Cell Biocatalysis with Organocatalysis Upgrades Alcohol Feedstocks in a Mild, Aqueous, One-Pot Process - Advances in Engineering

About the author

Kelsey Stewart joined the Colorado School of Mines as a Ph.D student in Applied Chemistry in August 2017, under the supervision Asst. Professor Dylan Domaille. Prior to that, she received a B.A. in Chemistry from Minot State University. While at Minot State, she worked on a project modifying epigenetic markers to induce differentiation in leukemic cell lines. Her current work focuses on interfacing chemical catalysts with living microbes to realize new products through biosynthetic means.

About the author

Emily Hicks was born and raised in Broomfield, Colorado and graduated from Broomfield High School in 2014. Wanting to stay in Colorado, she attended Colorado School of Mines in Golden. She joined the Domaille research group in fall 2017, assisting with the generation of an aldehyde-producing strain of bacteria used with biocompatible catalysts for biomass conversion applications. This is where she discovered her love of working in a lab. She graduated in 2018, with a B.S. in Chemical & Biochemical Engineering and a minor in Biomedical Engineering. After graduation she interned at Vitro Biopharma in Golden, gaining experience manufacturing stem cells and media for research and clinical use.

Now, she is a professional research assistant in the Skaggs School of Pharmacy and Pharmaceutical Sciences at Anschutz Medical Campus, studying the differences in drug metabolism between infants and adults in the Lampe Lab. When not in the lab, Emily enjoys hiking, camping, crafting, and spending time with her family, boyfriend, and cats.

About the author

Prof. Dylan Domaille received his PhD in chemistry in the chemical biology program from the University of California, Berkeley, where he worked with Chris Chang to develop small-molecule fluorescent sensors for tracking intracellular metal ions in living cells. He continued his studies with Jen Cha as a postdoctoral scholar in the Department of Chemical Engineering at the University of Colorado, Boulder, where he expanded his research to include M13 bacteriophage-based biosensors, stress-relaxing hydrogel 3D cell matrices, and biocompatible chemical processes.

Dylan joined the faculty in the Department of Chemistry at the Colorado School of Mines as an assistant professor in the fall of 2017 where his research focuses on questions in the fields bioenergy and biomaterials.

Reference

Kelsey N. Stewart, Emily G. Hicks, and Dylan W. Domaille. Merger of Whole Cell Biocatalysis with Organocatalysis Upgrades Alcohol Feedstocks in a Mild, Aqueous, One-Pot Process. ACS Sustainable Chemistry & Engineering 2020, volume 8, page 4114−4119.

Go To ACS Sustainable Chemistry & Engineering

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