Environmental Engineering Reference
In-Depth Information
corn fiber could become a valuable feedstock in the United States [31]. The use of
agricultural wastes is not without potential drawbacks, however. Most crop residues are
currently plowed into the soil to sustain soil quality by increasing the soil organic carbon
pool, enhancing activity of soil fauna, and minimizing soil erosion, and soil scientists caution
that diversion of waste biomass for fuel must be undertaken cautiously [32].
Softwood forest thinnings are also being explored as potential feedstocks. Lumber
manufacturing, timber harvesting, and thinning of forests to prevent wildfires generate a large
quantity of softwood residues that require environmentally sound and cost-effective methods
of disposal. Research in this area in the United States is currently focusing on dilute sulfuric
acid hydrolysis and SO
2
-steam explosion pretreatments, followed by fermentation by a
Saccharomyces cerevisiae mutant yeast adapted to the inhibitory extractives and lignin
degradation products present in the softwood hydrolysates [33]. Testing of recombinant
xylosefermenting yeasts is also planned, and investigation is underway by Kemestrie, Inc.
(Sherbrooke, QC, Canada) to identify high-value coproducts that may be derived from
softwoods, focusing on antioxidants and other extractives [3].
2.3. Cellulase Engineering
The second important area in which improvement is needed for the commercialization of
fuel ethanol is the conversion of lignocellulosic feedstock into the sugars to be fermented.
Most current work in this area concentrates on improvement of cellulase expression, activity,
and production efficiency, with the goal of reducing the cost and increasing the extent of
cellulose hydrolysis.
Cellulase cost is a critical limiting factor in lignocellulose feedstock preparation. Current
estimates of cellulase cost range from 3 0-50 cents per gallon of ethanol; a goal of 5 cents per
gallon of ethanol is envisioned [3]. Thus, a 10-fold improvement in specific activity,
production efficiency, or some combination thereof, is required.
Cellulase improvement in any of the following five critical areas could substantially
improve the feasibility of bioethanol commercialization: thermostability, acid tolerance (to
withstand pretreatment acidification), cellulose binding affinity, specific activity, and reduced
nonspecific binding to lignin [14]. While these features are theoretically approachable by
genetic engineering techniques, use of these techniques is presently limited by the incomplete
understanding of cellulase catalysis. A primary reason for this is that cellulose-cellulase
systems involve soluble enzymes working on insoluble substrates, which represents a
substantial increase in complexity from homogeneous enzyme-substrate systems. In addition,
the catalytic system involves the synergistic activities of three different enzymes [3]. Still, a
number of promising avenues are currently being explored.
2.3.1. Cellulase component engineering
. The most fundamental improvements that are
needed are within the cellulase components themselves, these are the endoglucanases,
exoglucanases, and cellobiohydrolases (or beta-glucosidases). Using Trichoderma,
Clostridium, Cellulomonas, and Thermobifida, among others, efforts are underway to
improve activity, expression, and specificity of these components through site-directed
mutagenesis, use of heterologous promoters to direct transcription, and modeling to reveal
structure-function relationships [34-39].
One of the most active cellulase components known is the endoglucanase E1 from
Acidothermus cellulolyticus. Two leading industrial enzyme producers, Novozymes
(www.novozymes.com) and Genencor International (www.genencor.com), are currently
