Carbon Conserving Microbial Pathway for the Production of Caprolactam from Renewable Feedstocks

When microorganisms use biological feedstocks such as sugars or glycerol to make more reduced chemicals, deoxygenation most often occurs via the loss of the feedstock’s carbon in the form of carbon dioxide (CO2). Feedstocks typically account for more than half of the total costs of a bioprocess, making them the most expensive component. Thus, the carbon lost as CO2 generally precludes profitability for many bio-based endeavors. To overcome this economics challenge, a unique carbon conserving (C2) platform technology is being developed at ZymoChem to minimize or eliminate CO2 production when microbes convert bio-based feedstocks into chemicals. ZymoChem’s C2 technology increases the maximum product yields by up to 50%, which could substantially decrease both capital costs and operating costs, in turn enabling the commercial-scale production of many chemicals in a sustainable, profitable manner. This Phase I SBIR project will focus on developing a novel biosynthetic pathway for producing ε-caprolactam from metabolic intermediates that can be made by microbes using ZymoChem’s C2 technology. One of two goals during Phase I will be to validate the feasibility of the proposed ε-caprolactam pathway. To accomplish this objective, in vitro studies will be first performed to identify highly functional enzymes for each pathway reaction. Next, the concerted functionality of all pathway reactions will be established in a microbial host for the in vivo production of caprolactam. The second goal of this Phase I project will be to develop our core C2 biosynthetic pathway in engineered strains of E. coli (available through the Technology Transfer Opportunity) that have improved anaerobic production of pyruvate, wherein the objective will be to have these strains improved our C2 technology, and specifically for the production of ε-caprolactam. Completing these objectives will represent the first example of the microbial production of ε-caprolactam, establishing the foundation for Phase II efforts at ZymoChem that will encompass strain engineering, pathway optimization, and scale-up R&D at the pilot level. ZymoChem’s new C2 approach to biologically synthesize chemicals while improving theoretical yields up to 50% establishes a means to substantially reduce production costs. Consequently, this economical benefit creates a probable opportunity to supplant petroleum-based state of the art and penetrate multiple markets in the chemical industry. With that said, commercialization of ZymoChem’s C2 technology for the production of ε-caprolactam will have several broad-reaching societal benefits. The renewable bioprocess with the company’s C2 technology will be cost-competitive with state-of-the-art methods – often an inherent limitation with most chemical- oriented biotechnologies – in turn providing an opportunity to penetrate ε-caprolactam’s ~$10 billion/yr market and displace the toxic and environmentally detrimental petroleum-based processes currently used to produce this value-added chemical. Commercialization of ZymoChem’s bioprocess for ε-caprolactam will also enable the production of bio-based nylon 6 (the polymer made from ε-caprolactam) and help “green” value chains across many sectors such as the textile, food, and automotive industries, which are highly reliant on using ε-caprolactam and nylon 6.