Plant derived feedstocks for industrial bioprocesses are made up of a complex set of materials. Carbohydrates in plants are used to store energy (e.g. starch) and provide structure (e.g. cellulose). These sugars can be in simple or complex forms, consisting of different sugar types linked together by a variety of chemical bonds, and at times decorated with additional chemical components.
This natural variety can make it difficult to break down and convert parts of these feedstocks, resulting in lost yield and productivity for producers. Mascoma’s CBP, addresses this problem in two ways. First, tailored enzyme systems are assembled into a robust yeast platform, and secreted to break down plant derived sugars into monomers at low marginal cost. Second, the released sugars (glucose, xylose, arabinose, mannose, galactose, etc.) are fermented to products like ethanol at high yield with minimal byproduct formation.
Tailored enzyme cocktails delivered at zero cost
Hemicellulose is a structural carbohydrate found in plant materials. It is a heterogeneous polymer of sugars, primarily xylan, with chemical decorations (acetyl and glucuronoyl groups among others) added to protect it from microbial attack. Typical processes to convert lignocellulose rely on either harsh acid treatment or expensive enzymes to break down hemicellulose to monomers. Hemicellulose that is not converted is extremely inhibitory to cellulases, reducing overall hydrolysis performance.
At Mascoma, we have developed a specialized enzyme suite to deal with hemicellulose that can be expressed in CBP strains. Enzymes from all classes known to hydrolyze carbohydrates, were developed for high level expression and produced in yeast, and then purified. These candidate enzymes were then tested for their ability to synergistically hydrolyze complex hemicellulose. Enzymes targeting all known bonds in the hemicellulose mixture were discovered. The data shown below demonstrates how a complex mix of 11 enzymes can reach over 90 percent hydrolysis yield at modest loadings (2.5 mg enzyme protein/g xylose present). This is in contrast to best hydrolysis obtained with “off the shelf” hemicellulase enzymes sourced from ENZCO, which provided at best 60 percent hydrolysis at a four times higher loading rate.
During fermentation, CBP strains produce enzyme cocktails as a small fraction of the cell mass that is normally accumulated as sugars are converted to ethanol. This means that these enzymes can be delivered to the fermentation at zero cost.
Specialized metabolic pathways for high rates, titers and yields
Xylose fermentation is not a capability that traditional ethanol producing yeast strains (Saccharomyces cerevisiae) possess. Achieving high rates, yields and titers under challenging industrial conditions has presented a barrier for ethanol producers. Mascoma has developed robust xylose fermentation capabilities by optimizing a recombinant xylose fermentation pathway. These strains of yeast have the ability to grow anaerobically on xylose at rates equivalent to those on glucose, to consume >95 percent of the xylose present, even under toxic conditions, and to ferment this xylose to ethanol at high yield. The figure below shows the performance of a xylose utilizing yeast biocatalyst for fermenting a xylose under two conditions: 1) a condition where xylose is the primary sugar present (in this case hemicellulose isolated from pretreated hardwood, and 2) a condition where xylose and glucose are present at approximately equal levels (in this case a hydrolysate generated from a pretreated agricultural waste).
Robust, high performance combinations
At Mascoma, we use advanced genetic tools to produce proprietary biocatalysts that combine the best metabolic pathways and enzyme cocktails for a particular application. These high performance combinations can create significant value for our customers across a wide range of industrial conditions.