Is There An Enzyme In Your Future?
The August 17 announcement, by Genencor International, Inc. (Rochester, NY) and five industrial partners, of a method for reducing the chemical steps in vitamin C synthesis from six to two, underscores the growing importance of enzymes in organic synthesis. Traditionally, vitamin C (ascorbic acid) production involves six chemical steps and one biotransformation (with the help of a microorganism) to form the product.
Fig. 1: Ascorbic Acid
After identifying enzymes that could carry out most of the chemistry, Genencor and its collaborators expressed all these proteins in a single gram-negative bacterium, fed the bug corn glucose, and out emerged 2-keto-L-gulonic acid. This product is then esterified and subjected to ring closure to produce the vitamin in very high yield and purity.
The five-year, $15.6 million Advanced Technology Program project was administered by the National Institute of Standards and Technology (NIST). Project team members and their contributions are:
- Genencor the joint venture administrator, supplying expertise in fermentation, reaction pathway engineering, high-performance microbial strains, and enzymology
- Eastman Chemical Co. chemical catalysis, separations, and process integration
- Electrosynthesis Co. electroprocessing and membrane technology for downstream processing
- Microgenomics, Inc. genetics know-how, identification of synthesizing enzymes
- Argonne National Laboratory functional genomics, downstream processing
Genencor, which holds more than 1,200 worldwide patents, exploits biodiversity, genomics/biotechnology, and reaction pathway engineering to achieve three goals: discovering and identifying industrial-grade enzymes; improving those enzymes and expressing them in hardy microorganisms; and redesigning synthetic chemical pathways to exploit their other technologies.
The $600 million world market for the vitamin has recently experienced wild price fluctuations, to levels as low as $5/kg. So, as chiral bioactive materials go, it's safe to say ascorbic acid is already a commodity. Still, according to Richard LaDuca, Genencor's director of technology and business development, either Genencor or a licensee will eventually make a business out of this process because of the tremendous cost savings, both in physical plant and processing. LaDuca estimates that an enzyme-based ascorbic acid plant would cost about half as much to build as today's plants that make the vitamin using mostly conventional chemistry. And even at this early stage, pound for pound, the multiple enzyme pathway is cost-competitive. "Both the number and sizes of reactors are smaller with our process, so we're in essence shrinking the size of the factory," LaDuca said. "Many variables will determine how our cost-competitiveness eventually plays out, but since we are committed to investing in this area for another two years, we're confident that the economics are going to get even better."
Making vitamin C per se is not terribly exciting, but stands as a proof of concept for an exciting new manufacturing idea: carrying out multiple reactions in one microorganism. "We're seeking to identify and use families of enzymes capable of performing many types of organic transformations," LaDuca told BioResearch Online. "In that regard our technology is very much generic. As we build a pathway to ascorbic acid, we have the ability to stop the reaction at any point or to add other enzymes to divert the synthesis towards other products. The pathway we're building has the general capability for producing chemical synthons for other value-added products."
Genencor has based a successful business on industrial enzymes, and now is seeking to couple that expertise with reaction pathway engineering. Their success may determine how easily enzymes are accepted by chemical and pharmaceutical manufacturers. One company goal, for example, is to understand the interplay between upstream (mostly synthetic) and downstream (purification) unit operations, and to design a manufacturing process so that both stages are optimized.
Multiple enzyme hosts are critical to Genencor's success. Manufacturers are reluctant to construct a bioprocessing plant for one reaction out of eight in a complex synthesisthey will simply find another way or redesign their process. A bioprocess that carried out the equivalent of six out of eight steps, however, would be very attractive. In addition, large-scale manufacturing should be continuous or semi-continuous, whereas most bioprocesses operate in batch mode.
Many chemists believe that one-day enzymes will carry out a wide range of industrially significant organic reactions, and that these syntheses will occur outside a microbiological host. LaDuca envisions "being able to go to a refrigerator full of enzymes and use them in chemical synthesis and manufacturing the way we use conventional reagents today. I'm not painting a picture of a world where enzymes perform all chemical reactions. As a realist, I'd settle for a world where chemists use the best chemical and enzyme-based processes."
For more information: Richard LaDuca, Director of Technology and Business Development, Genencor International, Inc., 925 Page Mill Rd., Palo Alto, CA 94304. Tel: 650-846-7500.