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Our research interests are in biochemical engineering, with particular emphasis on biocatalysis, biomaterials, and bioenergy. Past projects in our research group include the structural characterization and activation of enzymes in non-aqueous reaction media, with the goal of overcoming process limitations imposed by aqueous conditions; the development of enzyme- and cell-based microscale systems for high-throughput bioactivity screening and drug discovery; and sustainable production of biofuels from lignocellulosic feedstocks. Current research projects expand on these themes, combining new experimental methods and multiphysics models to advance fundamental understanding and practical applications of extremophilic organisms, artificial metalloenzymes, and biological energy systems. Three major current research topics are described below.
Mechanistic studies of adaptation to harsh environments by extremophilic microorganisms
Understanding the survival mechanisms by which cells adapt to extreme conditions is of fundamental interest in microbiology. Furthermore, the transfer of favourable physiological characteristics across microbial species has important implications for microbial ecology as well as the development of robust microbial hosts for the production of various bioproducts. In particular, enhancing the tolerance of microorganisms to stressful environments; for example, elevated temperatures and alcohol/solvent concentrations, holds enormous promise for biotechnology. In this connection, the gamma-prefoldin (γPFD), a filamentous chaperone from the deep-sea, hyperthermophilic methanogen Methanocaldococcus jannaschii, may prove particularly valuable.
Our previous research has focused on utilizing the γPFD as a template for enzyme immobilization and conductive nanowires, as a central component of protein-based hydrogels, and as a protein building block for the assembly of elaborate 2D nanostructures. Currently we are investigating 1) the specific mechanism(s) by which γPFD protects microbial cells against harsh environments, and 2) whether enhanced thermotolerance conferred by filamentous PFDs is a general adaptation used by archaea to survive extreme temperatures. Determining the functions of filamentous PFDs in the survival of extremophilic archaea will advance our understanding of life in extremis, and may point the way toward conferring greater tolerance to environmental extremes across microbial species and cell factories for biotechnology applications.
Electronically conductive nanowire comprised of c3-type cytochromes conjugated along the longitudinal axis of the filamentous gamma-prefoldin. From Chen et al., ACS Nano 2020 14 (6), 6559-6569. DOI: 10.1021/acsnano.9b09405
Structure-function relationships and biosynthetic applications of artificial metalloenzymes
Complex molecules produced in microorganisms by biosynthesis are the inspiration or the actual molecules in products ranging from medicines to agrochemicals. Synthetic biology enables the preparation of many complex molecules by incorporating into a suitable production strain biosynthetic genes obtained from organisms that are rare or difficult to cultivate in a laboratory. Although natural organisms produce molecules with complex structures that are the envy of chemists, the diversity of these molecules is limited by the scope of chemical reactions catalyzed by natural enzymes.
Recently, artificial metalloenzymes have been created that catalyze a series of abiotic reactions, including group transfers to olefins and C-H bonds, cross-coupling, asymmetric hydrogenation, and ring-closing olefin metathesis. If artificial enzymes could be incorporated into biosynthetic pathways, then microorganisms could produce a broader range of products by a combination of nature’s reactions and chemists’ imaginations. Current research in collaboration with the Hartwig and Keasling groups will expand the repertoire of unnatural products that can be the leads for medicines, new agrochemicals, and new monomers for polymerization, along with molecules for unforeseen applications, by combining the capabilities of synthetic biology to produce natural and unnatural products in engineered microorganisms with the wide range of abiotic reactions of artificial metalloenzymes.
Structure of Fe-CYP119 showing active-site residues modified during directed evolution of the corresponding artificial metalloenzyme containing iridium in place of iron to increase activity and selectivity for carbene insertions into C–H bonds. From Dydio et al., Science 2016 354 (6308), 102-106. DOI: 10.1126/science.aah4427
Sustainable production of biofuels and value-added chemicals from carbon dioxide
Biotechnology has a key role to play in developing a circular carbon economy as demand for industrial enzymes and biotherapeutics grows and biochemical processes displace fossil-based production of fuels, plastics, and commodity chemicals. Electromicrobial production processes, in which electricity or electrochemically-derived mediator molecules serve as energy sources to drive biochemical processes, represent an attractive strategy for the conversion of CO2 into carbon-based products. However, these systems have yet to be employed on an industrial scale, limiting our understanding of their potential performance and environmental benefits/impacts. We are currently part of a collaborative effort between UC Berkeley and GE Research that aims to develop an integrated, three-step process to directly capture and convert carbon dioxide (CO2) from ambient air into butanol, a platform molecule for diesel and jet fuels. Successful development of the proposed process may ultimately result in reduced energy imports and reduced greenhouse gas emissions by providing a clean alternative route towards producing fuels.
Reactor overview and formatotrophic growth strategies for conversion of CO2 into carbon-based products by microbial electrosynthesis. From Abel and Clark, ChemSusChem 2021, 14, 344–355. doi.org/10.1002/cssc.202002079
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