Bacteria and Cellulosic Biofuels

Claire Durkin
November 29, 2011

Submitted as coursework for PH240, Stanford University, Fall 2011


Fossil fuels are a non-renewable source of energy. Because finite amounts of coal, petroleum, and natural gas deposits remain on Earth, a shift toward sustainable energy production is required to maintain the world energy demand. Based on current fuel consumption rates, it is estimated that oil reserves will be depleted between 2062 and 2094. [1]

Additionally, American dependence on foreign oil reserves has increased over the past half-century with serious political implications. The U.S. currently imports 59 percent of its oil supplies, up from 21.5 percent in 1970. [2] A shift toward energy autonomy would decrease policy limitations set by other countries and allow greater American independence from oil producing nations.

The 2007 Energy Independence and Security Act recognized the need for domestic, renewable energy resources, requiring the production of 36 billion gallons of renewable fuels by 2022. This represents a fivefold increase from current production levels, leading many in science and industry to look toward biofuels for energy in the coming years. [3]

Cellulosic vs. Grain Biofuel

Two main sources of plant matter are used for biofuels: grains and cellulose. Grains are the most common source for biofuel production. [4] Because they are comprised of starches and sugars, grains can be converted into ethanol efficiently and for relatively low costs. In contrast, cellulose must be extracted from the tough plant cell walls and converted into sugar before it can used for energy production, a more involved and energy intensive process than grain conversion. [4] Production costs of starch-to-ethanol plants is $0.88 per gallon compared to $1.50 for cellulose-to-ethanol conversion. [5]

While production of biofuels from grains currently is cheaper than from cellulose, studies indicate that producing ethanol from cellulose is better for the environment than from grains. Corn is the most widespread source of ethanol in the United States. Comparing grams of carbon dioxide released per megajoule of energy in fuel, corn's use in fuel production reduces greenhouse gas emissions 20% from gasoline, while cellulosic biofuels reduce emissions 70%. Corn ethanol also shows a greater carbon intensity than cellulosic ethanol when taking into account the indirect land use change impacts of biofuels (ILUC), which estimates changes in carbon emissions changes due to cropland alterations. [6]

Cellulose also has potential for being an important fuel source because it is a waste material, while grains have many alternate uses such as food supply. Cellulose is a polysaccharide found in the cell wall of all plants. [7] An abundant resource, cellulose often is harvested from switch grass, wood, straw, and stalks of corn stover. Paper, paperboard, food wastes, textiles, yard wastes, wood, and miscellaneous organics comprise about 72% of municipal solid waste by weight, which translates to about 130 million tons of cellulose-containing materials being sent to landfills every year from U.S. municipal trash. [8] Because cellulose cannot be digested by humans and is commonly discarded as food or paper waste-products, it is an inexpensive and widely available, making it ideal for fuel production. Additionally, scientists estimate that cellulose may yield about 3 times the energy as grain per unit weight. [9]

Inefficiency of Current Cellulosic Biofuel Production Methods

Ethanol is made from cellulosic plant mass with two common approaches, cellulolysis and gasification. Cellulolysis is a biochemical reaction that involves pretreatment, enzymatic conversion of cellulose to sugar or starch, separation of sugars from plant residues, fermentation of sugar using microbes, distillation, and dehydration. [10] Gasification is a thermochemical reaction in which dried plant matter is burned into synthesis gas, which then reacts with metal catalysts to form ethanol or other hydrocarbons. This process, called Fischer-Tropsch synthesis, is not economically competitive with fossil fuels. Production costs are $9/GJ compared to $5/GJ for diesel. [11] At these current prices, cellulosic biofuel is not a competitive product. There is a definite need for a cheaper and more efficient conversion processes.

Engineering Bacteria to Produce Biodiesel

Bacterial reactors designed with synthetic biology are a promising approach for making competitively viable biofuels. In synthetic biology, metabolic pathways in bacteria are engineered to break down cellulose into desired hydrocarbon fuel compounds. Optimization methods aim to increase production yield efficiency, while ensuring that cells obtain enough nutrients to proliferate.

Dr. Jay Keasling's synthetic biology lab at UC Berkeley recently engineered E. coli to produce ready-to-use diesel directly from cellulosic biomass. E. coli was chosen because it is a well-studied model organism and reproduces rapidly compared to other bacteria.

First, hemicellulase genes from Clostridium stercorarium and Bacteroides ovatus were cloned into the E. coli and a short amino acid sequence tag was used so that resulting enzymes would be secreted. These genes express hemicellulase, which breaks down cellulose into sugars.

Second, E. coli was engineered to produce structurally tailored fatty esters (diesel), alcohols, and waxes from sugar. Thioesterase-catalyzed hydrolysis of fatty acyl-ACP was combined with fatty acyl-CoA-synthase-catalyzed reactivation of the fatty acid carboxylate group. The thioesterase-catalyzed hydrolysis was energetically favorable, overproducing fatty acids and deregulating fatty acid biosynthesis. Fatty acid metabolism was diverted to fatty acyl-CoA, which is an important substrate for production of alcohols and esters.The Keasling Lab's successful conversion of cellulose directly to biodiesel represents a step toward engineering synthetic bacteria to produce renewable fuels. However, biodiesel yields are only at approximately 10% of the maximum theoretical conversion, so much work needs to be done before scale-up and realization of a commercial product. [12,13]


The United States is reliant on foreign, non-renewable fuels at present. The enactment of the Energy Independence and Security Act of 2007 recognized the need for sustainable, domestic energy production. Cellulose is a promising source for biofuels because it is cheap and abundant, but current technologies for cellulose conversion to energy are too expensive to realistically compete on the market. Engineering bacteria metabolisms through synthetic biology to efficiently convert cellulose directly to biodiesel is a possible solution to reduce production costs.

© Claire Durkin. The author grants permission to copy, distribute and display this work in unaltered form, with attribution to the author, for noncommercial purposes only. All other rights, including commercial rights, are reserved to the author.


[1] C. Wessner, "FYI: What Is the Best Guess as to When the World's Oil Reserves Will Run Out?" Popular Science, February 1999, p. 76.

[2] I. Rutledge, Addicted to Oil: America's Relentless Drive for Energy Security. (I. B. Tauris, 2005).

[3] J. M. Broder, "Bush Signs Broad Energy Bill," New York Times, 19 Dec 07.

[4] R. B. Gupta and A. Demirbas, Gasoline, Diesel, and Ethanol Biofuels from Grasses and Plants (Cambridge, 2010).

[5] A. McAloon et al., "Determining the Cost of Producing Ethanol from Corn Starch and Lignocellulosic Feedstocks," U.S. National Renewable Energy laboratory, NREL/TP-580-22893, October 2000.

[6] T. Searchinger et al., "Use of U.S. Croplands for Biofuels Increases Greenhouse Gases Through Emissions from Land-Use Change," Science 319, 1238 (2008).

[7] R. M. Brown, Jr. and I. M. Saxena, eds., Cellulose: Molecular and Structural Biology (Springer, 2007).

[8] S. R. Qasim and W. Chiang, Sanitary Landfill Leachate: Generation, Control, and Treatment, (CRC Press, 1994).

[9] L. R. Lynd, "Overview and Evaluation of Fuel Ethanol from Cellulosic Biomass: Technology, Economics, the Environment, and Policy," Annu. Rev. Energy Environ. 21, 403 (1996).

[10] J. Y. Zhu et al., "Sulfite Pretreatment (SPORL) for Robust Enzymatic Saccharification of Spruce and Red Pine," Bioresource Technol. 100, 2411 (2009).

[11] M. Tijmensen, "Exploration of the Possibilities for Production of Fischer Tropsch Liquids and Power via Biomass Gasification," Biomass and Bioenergy 23, 129 (2002).

[12] D. Biello, "Bacteria Transformed into Biofuel Refineries," Scientific American, 27 Jan 10.

[13] E. J. Steen et al., "Microbial Production of Fatty-Acid-Derived Fuels and Chemicals from Plant Biomass," Nature 463, 559 (2010).