Perspectives and new directions for the production of bioethanol using consolidated bioprocessing of lignocellulose

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The U.S. DOE Energy Independence and Security Act (EISA) mandated attainment of a national production level of 36 billion gallons of biofuels (to be added to gasoline) by 2022, of which 21 billion gallons must be derived from renewable/sustainable feedstocks (e.g. lignocellulose). In order to attain these goals, the development of cost effective process technologies that can convert plant biomass to fermentable sugars must occur. An alternative route to production of bioethanol is the utilization of microorganisms that can both convert biomass to fermentable sugars and ferment the resultant sugars to ethanol in a process known as consolidated bioprocessing (CBP). Although various economic benefits and technology hurdles must be weighed in the course of choosing the CBP strategy to be pursued, we present arguments for developing the powerfully cellulolytic fungus, Trichoderma reesei, as an effective CBP microorganism.

Introduction

Biofuel production processes today are based on the fermentation of biomass sugars and must be improved to meet EISA goals [1]. These processes employ various configurations comprising the following unit operations: biomass collection and storage, comminution (size reduction), thermal chemical pretreatment, saccharification, fermentation, and product recovery. Recent technoeconomic analyses of the corn stover to ethanol process, currently based on dilute sulfuric acid and SSF (simultaneous saccharification and fermentation) (Figure 1a–c), report that the capital cost of pretreatment and production and/or purchase of cellulase enzymes remain the dominant cost hurdles to overcome [2, 3]. However, this process development history is dominated by conversion processes that require fermentable sugars for a subsequent process step, fermentation by either ethanologenic yeast or bacteria.

Some options proposed to reduce the cost of the conversion of lignocellulose to ethanol, include: eliminating pretreatment, increasing cellulose hydrolysis yield, enhancing the enzyme activity to reduce its consumption, and improving the fermentation process both in yield and specificity. CBP combines simultaneous saccharification of lignocellulose with fermentation of the resulting sugars into a single process step mediated by a single microorganism or microbial consortium. Compared to other less highly integrated configurations, CBP is distinctive because it does not contain a separate and dedicated process step for cellulase production [4••]. This factor alone helps lower the costs of converting lignocellulose to ethanol that is not yet economical owing to the high costs of biomass pretreatment and enzymatic saccharification. Evidence is accumulating that suggests that CBP may be feasible [5]. Ever since the concept of CBP was proposed in 1996, CBP research has focused on the development of new and evermore effective CBP microorganisms, which has been a key challenge [4••]. Bacteria and yeast have been the primary candidates for CBP research and some progress has been made in this regard [6••]. However, fungi have not been widely proposed as CBP microorganisms nor has progress been reported in regard to developing a fungal CBP candidate using genetic approaches. We will introduce the concept of traditional CBP briefly, and will then focus on a discussion of the fungal CBP concept. We will especially discuss the key properties of fungi relevant to CBP, our perceptions of the feasibility of fungal CBP, and then possible strategies for developing fungi for CBP and their potential application to industrial processes.

Section snippets

Development of bacterial and yeast CBP—brief progress and existing problems

Although no natural microorganism possesses all properties of lignocellulose utilization and ethanol production desired for CBP, some bacteria and fungi exhibit some of the needed traits (see Table 1). Traditionally, proponents of CBP processes have identified two primary developmental pathways capable of producing industrially viable CBP microbial strains. These are category I, engineering a cellulase producer, such as Clostridium thermocellum, to be ethanologenic; and category II, engineering

Properties of T. reesei relevant to CBP

Despite recent advances in engineering cellulases to be more efficient and less costly, the complete saccharification of pretreated lignocellulose still requires a very long time for digestion and high loadings of enzyme (30–50 mg enzyme per g of crystalline cellulose). Thus, a biorefinery consuming 10 000 tons of biomass per day will require many tons of cellulase preparation to operate. Today, only fungi naturally produce the needed titers of cellulase to meet this need. For example, T. reesei

Primary putative T. reesei genes required for the conversion of the lignocellulosic sugars to ethanol: strategies to improve ethanol production

The key T. reesei metabolic pathways for converting lignocellulosic sugars to ethanol include the glycolysis, pentose phosphate pathway, and ethanolic fermentation. From our systematic survey of the incomplete genome sequence of T. reesei (http://genome.jgi-psf.org/Trire2/Trire2.home.html), we identified virtually all genes involved in glycolysis, pentose phosphate pathway, xylose and arabinose assimilation pathways, and the classical ethanolic fermentation. We have demonstrated that T. reesei

Conclusion

CBP has the potential to reduce the cost of the biological conversion of lignocellulose to ethanol significantly, and accumulating data are demonstrating its feasibility. Though some progress has been made in both categories I and II CBP development, the limited ability of the latter system for producing enzymes in sufficient quantity and quality for lignocellulose degradation remains a challenge. It was initially expected that this problem could be overcome by introduction of heterologous

References and recommended reading

Papers of particular interest, published within the period of review, have been highlighted as:

  • • of special interest

  • •• of outstanding interest

Acknowledgement

This work was supported by the Laboratory Directed Research and Development (LDRD) Program at the National Renewable Energy Laboratory.

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