Â鶹ÒùÔº

Building on their success developing a cleaner way to make valuable organic acids, researchers from the Center for Advanced Bioenergy and Bioproducts Innovation (CABBI) have pushed one product closer to commercialization with a major upgrade in yield.

A CABBI team from the University of Illinois Urbana-Champaign and Princeton University re-engineered the metabolism of the yeast Issatchenkia orientalis to supercharge its fermentation of plant glucose into succinic acid—an important industrial chemical used in food additives and a diverse array of agricultural and pharmaceutical products. Since I. orientalis can tolerate highly acidic conditions, it's an ideal host for organic acid production.

This natural fermentation process, relying on yeasts and renewable plant material, is more environmentally friendly than conventional production using petrochemicals. But cost remains a barrier to commercial adoption, according to CABBI Conversion Theme Leader Huimin Zhao, Professor of Chemical and Biomolecular Engineering (ChBE) at Illinois and a lead author on the study.

Improving the yield—the amount of succinic acid produced from a gram of glucose—would make the process more economically viable.

The CABBI team previously created a cost-effective, end-to-end succinic acid pipeline using an engineered strain of I. orientalis, with added enzymes and genes that allowed it to better utilize sucrose from sugarcane and boost its production of succinic acid.

That pilot project showed the engineered yeast could produce 110 grams per liter (g/L) of succinic acid in low pH conditions, with an overall yield of 64% after fermentation and processing.

In the new study, published in , they used a different metabolic engineering strategy on I. orientalis called "decompartmentalization" and boosted yield to 85%—a significant improvement, Zhao said.

All living cells, including the yeast cells used in this study, use energy from sugars like glucose to make NADH (nicotinamide adenine dinucleotide plus hydrogen), a coenzyme that helps certain enzymes perform chemical reactions.

NADH acts like a rechargeable battery storing energy. This energy can be used to build succinic acid and other reduced chemicals—those gaining an electron through a reaction known as oxidation-reduction.

However, there may not be enough NADH in the right part of the cell to support production. This problem is especially challenging in yeasts because their cells contain different compartments that do different jobs—such as mitochondria for energy production and peroxisomes for the breakdown of certain chemicals—and NADH is spread out across these different compartments.

To overcome this, researchers moved key parts of the NADH-making system from the mitochondria into the main area of the cell, the cytosol, where succinic acid is made. This "decompartmentalization" gave the cells more energy-producing NADH where it was needed, further improving succinic acid production from renewable sugars.

"These advances bring us closer to greener manufacturing processes that benefit both the environment and the economy," said Vinh Tran, primary author on the paper and former Ph.D. student with Zhao in ChBE and at the Carl R. Woese Institute for Genomic Biology (IGB).

Succinic acid is one of the U.S. Department of Energy's top 12 bio-based value-added chemicals, and a well-engineered microbial strain is highly sought after for its efficient and economical production.

The CABBI team's earlier analysis showed that the minimum product selling price (MPSP) is most sensitive to fermentation yield, indicating that further yield improvement was essential for enhancing the financial viability of the engineered I. orientalis.

Boosting succinic acid yield to 0.85 grams per gram of glucose (g/g) reduced the MPSP to 97 cents per kilogram, down from $1.30 previously.

"That is huge," Zhao said. "For those industrial chemicals, even a few cents is a big reduction."

About 700,000 tons of succinic acid are produced each year, a market totaling $200 million, so the savings quickly multiply, Zhao said.

Companies have already expressed interest in scaling up this technology for commercial use, which could potentially happen within a year, Zhao said.

The CABBI team is applying these approaches to produce other industrially important organic acids, such as 3-hydroxypropionic acid (3-HP), as part of its work to develop biofuels and biochemicals from biomass crops.