New advanced biofuel identified as an alternative to diesel fuel

ScienceDaily (Sep. 28, 2011) — Researchers with the U.S Department of Energy (DOE)'s Joint BioEnergy Institute (JBEI) have identified a potential new advanced biofuel that could replace today's standard fuel for diesel engines but would be clean, green, renewable and produced in the United States.

Using the tools of synthetic biology, a JBEI research team engineered strains of two microbes, a bacteria and a yeast, to produce a precursor to bisabolane, a member of the terpene class of chemical compounds that are found in plants and used in fragrances and flavorings. Preliminary tests by the team showed that bisabolane's properties make it a promising biosynthetic alternative to Number 2 (D2) diesel fuel.

"This is the first report of bisabolane as a biosynthetic alternative to D2 diesel, and the first microbial overproduction of bisabolene in Escherichia coli and Saccharomyces cerevisiae," says Taek Soon Lee, who directs JBEI's metabolic engineering program and is a project scientist with Lawrence Berkeley National Laboratory (Berkeley Lab)'s Physical Biosciences Division. "This work is also a proof-of-principle for advanced biofuels research in that we've shown that we can design a biofuel target, evaluate this fuel target, and produce the fuel with microbes that we've engineered."

Lee is the corresponding author of a paper reporting this research in the journal Nature Communications. Co-authoring this paper were Pamela Peralta-Yahya, Mario Ouellet, Rossana Chan, Aindrila Mukhopadhyay and Jay Keasling.

The rising costs and growing dependence upon foreign sources of petroleum-based fuels, coupled with scientific fears over how the burning of these fuels impacts global climate, are driving the search for carbon-neutral renewable alternatives. Advanced biofuels -- liquid transportation fuels derived from the cellulosic biomass of perennial grasses and other non-food plants, as well as from agricultural waste -- are highly touted for their potential to replace gasoline, diesel and jet fuels. Unlike ethanol, which can only be used in limited amounts in gasoline engines and can't be used at all in diesel or jet engines, plus would corrode existing oil pipelines and tanks, advanced biofuels are drop-in fuels compatible with today's engines, and delivery and storage infrastructures.

"We desperately need drop-in, renewable biofuels that can directly replace petroleum-derived fuels, particularly for vehicles that cannot be electrified," says co-author Keasling, CEO of JBEI and a leading authority on advanced biofuels. "The technology we describe in our Nature Communications paper is a significant advance in that direction."

JBEI is one of three Bioenergy Research Centers established by the DOE's Office of Science in 2007. Researchers at JBEI are pursuing the fundamental science needed to make production of advanced biofuels cost-effective on a national scale. One of the avenues being explored is sesquiterpenes, terpene compounds that contain 15 carbon atoms (diesel fuel typically contains 10 to 24 carbon atoms).

"Sesquiterpenes have a high-energy content and physicochemical properties similar to diesel and jet fuels," Lee says. "Although plants are the natural source of terpene compounds, engineered microbial platforms would be the most convenient and cost-effective approach for large-scale production of advanced biofuels."

In earlier work, Lee and his group engineered a new mevalonate pathway (a metabolic reaction critical to biosynthesis) in both E. coli and S. cerevisiae that resulted in these two microorganisms over-producing a chemical compound called farnesyl diphosphate (FPP), which can be treated with enzymes to synthesize a desired terpene. In this latest work, Lee and his group used that mevalonate pathway to create bisabolene, which is a precursor to bisabolane.

"We proposed that the generality of the microbial FPP overproduction platforms would allow for the biosynthesis of sesquiterpenes," Lee says. "Through multiple rounds of large-scale preparation in shake flasks, we were able to prepare approximately 20 milliliters of biosynthetic bisabolene, which we then hydrogenated to produce bisabolane."

When they began this work, Lee and his colleagues did not know whether bisabolane could be used as a biofuel, but they targeted it on the basis of its chemical structure. Their first step was to perform fuel property tests on commercially available bisabolene, which comes as part of a mixture of compounds. Convinced they were onto something, the researchers then used biosynthesis to extract pure biosynthetic bisabolene from microbial cultures for hydrogenation into bisabolane. Subsequent fuel property tests on the bisabolane were again promising.

"Bisabolane has properties almost identical to D2 diesel but its branched and cyclic chemical structure gives it much lower freezing and cloud points, which should be advantageous for use as a fuel," Lee says. "Once we confirmed that bisabolane could be a good fuel, we designed a mevalonate pathway to produce the precursor, bisabolene. This was basically the same platform used to produce the anti-malarial drug artemisinin except that we introduced a terpene synthase and further engineered the pathway to improve the bisabolene yield both in E. coli and yeast."

Lee and his colleagues are now preparing to make gallons of bisabolane for tests in actual diesel engines, using the new fermentation facilities at Berkeley Lab's Advanced Biofuels Process Demonstration Unit. The ABPDU is a 15,000 square-foot state-of-the art facility, located in Emeryville, California, designed to help expedite the commercialization of advanced next-generation biofuels by providing industry-scale test beds for discoveries made in the laboratory.

"Once the complete fuel properties of hydrogenated biosynthetic bisabolene can be obtained, we'll be able to do an economic analysis that takes into consideration production variables such as the cost and type of feedstock, biomass depolymerization method, and the microbial yield of biofuel," Lee says. "We will also be able to estimate the impact of byproducts present in the hydrogenated commercial bisabolene, such as farnesane and aromatized bisabolene."

Ultimately, Lee and his colleagues would like to replace the chemical processing step of bisabolene hydrogenation with an alkene reductase enzyme engineered into the E. coli and yeast so that all of the chemistry is performed within the microbes.

"Enzymatic hydrogenation of this type of molecule is a very challenging project and will be a long term goal," Lee says. "Our near-term goal is to develop strains of E. coli and yeast for use in commercial-scale fermenters. Also, we will be investigating the use of sugars from biomass as a source of carbon for producing bisabolene."

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The above story is reprinted (with editorial adaptations by ScienceDaily staff) from materials provided by DOE/Lawrence Berkeley National Laboratory.

Journal Reference:

Pamela P. Peralta-Yahya, Mario Ouellet, Rossana Chan, Aindrila Mukhopadhyay, Jay D. Keasling, Taek Soon Lee. Identification and microbial production of a terpene-based advanced biofuel. Nature Communications, 2011; 2: 483 DOI: 10.1038/ncomms1494

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Disclaimer: Views expressed in this article do not necessarily reflect those of ScienceDaily or its staff.

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New advanced biofuel identified as an alternative to diesel fuel

ScienceDaily (Sep. 28, 2011) — Researchers with the U.S Department of Energy (DOE)'s Joint BioEnergy Institute (JBEI) have identified a potential new advanced biofuel that could replace today's standard fuel for diesel engines but would be clean, green, renewable and produced in the United States.

Using the tools of synthetic biology, a JBEI research team engineered strains of two microbes, a bacteria and a yeast, to produce a precursor to bisabolane, a member of the terpene class of chemical compounds that are found in plants and used in fragrances and flavorings. Preliminary tests by the team showed that bisabolane's properties make it a promising biosynthetic alternative to Number 2 (D2) diesel fuel.

"This is the first report of bisabolane as a biosynthetic alternative to D2 diesel, and the first microbial overproduction of bisabolene in Escherichia coli and Saccharomyces cerevisiae," says Taek Soon Lee, who directs JBEI's metabolic engineering program and is a project scientist with Lawrence Berkeley National Laboratory (Berkeley Lab)'s Physical Biosciences Division. "This work is also a proof-of-principle for advanced biofuels research in that we've shown that we can design a biofuel target, evaluate this fuel target, and produce the fuel with microbes that we've engineered."

Lee is the corresponding author of a paper reporting this research in the journal Nature Communications. Co-authoring this paper were Pamela Peralta-Yahya, Mario Ouellet, Rossana Chan, Aindrila Mukhopadhyay and Jay Keasling.

The rising costs and growing dependence upon foreign sources of petroleum-based fuels, coupled with scientific fears over how the burning of these fuels impacts global climate, are driving the search for carbon-neutral renewable alternatives. Advanced biofuels -- liquid transportation fuels derived from the cellulosic biomass of perennial grasses and other non-food plants, as well as from agricultural waste -- are highly touted for their potential to replace gasoline, diesel and jet fuels. Unlike ethanol, which can only be used in limited amounts in gasoline engines and can't be used at all in diesel or jet engines, plus would corrode existing oil pipelines and tanks, advanced biofuels are drop-in fuels compatible with today's engines, and delivery and storage infrastructures.

"We desperately need drop-in, renewable biofuels that can directly replace petroleum-derived fuels, particularly for vehicles that cannot be electrified," says co-author Keasling, CEO of JBEI and a leading authority on advanced biofuels. "The technology we describe in our Nature Communications paper is a significant advance in that direction."

JBEI is one of three Bioenergy Research Centers established by the DOE's Office of Science in 2007. Researchers at JBEI are pursuing the fundamental science needed to make production of advanced biofuels cost-effective on a national scale. One of the avenues being explored is sesquiterpenes, terpene compounds that contain 15 carbon atoms (diesel fuel typically contains 10 to 24 carbon atoms).

"Sesquiterpenes have a high-energy content and physicochemical properties similar to diesel and jet fuels," Lee says. "Although plants are the natural source of terpene compounds, engineered microbial platforms would be the most convenient and cost-effective approach for large-scale production of advanced biofuels."

In earlier work, Lee and his group engineered a new mevalonate pathway (a metabolic reaction critical to biosynthesis) in both E. coli and S. cerevisiae that resulted in these two microorganisms over-producing a chemical compound called farnesyl diphosphate (FPP), which can be treated with enzymes to synthesize a desired terpene. In this latest work, Lee and his group used that mevalonate pathway to create bisabolene, which is a precursor to bisabolane.

"We proposed that the generality of the microbial FPP overproduction platforms would allow for the biosynthesis of sesquiterpenes," Lee says. "Through multiple rounds of large-scale preparation in shake flasks, we were able to prepare approximately 20 milliliters of biosynthetic bisabolene, which we then hydrogenated to produce bisabolane."

When they began this work, Lee and his colleagues did not know whether bisabolane could be used as a biofuel, but they targeted it on the basis of its chemical structure. Their first step was to perform fuel property tests on commercially available bisabolene, which comes as part of a mixture of compounds. Convinced they were onto something, the researchers then used biosynthesis to extract pure biosynthetic bisabolene from microbial cultures for hydrogenation into bisabolane. Subsequent fuel property tests on the bisabolane were again promising.

"Bisabolane has properties almost identical to D2 diesel but its branched and cyclic chemical structure gives it much lower freezing and cloud points, which should be advantageous for use as a fuel," Lee says. "Once we confirmed that bisabolane could be a good fuel, we designed a mevalonate pathway to produce the precursor, bisabolene. This was basically the same platform used to produce the anti-malarial drug artemisinin except that we introduced a terpene synthase and further engineered the pathway to improve the bisabolene yield both in E. coli and yeast."

Lee and his colleagues are now preparing to make gallons of bisabolane for tests in actual diesel engines, using the new fermentation facilities at Berkeley Lab's Advanced Biofuels Process Demonstration Unit. The ABPDU is a 15,000 square-foot state-of-the art facility, located in Emeryville, California, designed to help expedite the commercialization of advanced next-generation biofuels by providing industry-scale test beds for discoveries made in the laboratory.

"Once the complete fuel properties of hydrogenated biosynthetic bisabolene can be obtained, we'll be able to do an economic analysis that takes into consideration production variables such as the cost and type of feedstock, biomass depolymerization method, and the microbial yield of biofuel," Lee says. "We will also be able to estimate the impact of byproducts present in the hydrogenated commercial bisabolene, such as farnesane and aromatized bisabolene."

Ultimately, Lee and his colleagues would like to replace the chemical processing step of bisabolene hydrogenation with an alkene reductase enzyme engineered into the E. coli and yeast so that all of the chemistry is performed within the microbes.

"Enzymatic hydrogenation of this type of molecule is a very challenging project and will be a long term goal," Lee says. "Our near-term goal is to develop strains of E. coli and yeast for use in commercial-scale fermenters. Also, we will be investigating the use of sugars from biomass as a source of carbon for producing bisabolene."

Recommend this story on Facebook, Twitter,
and Google +1:

Other bookmarking and sharing tools:

Story Source:

The above story is reprinted (with editorial adaptations by ScienceDaily staff) from materials provided by DOE/Lawrence Berkeley National Laboratory.

Journal Reference:

Pamela P. Peralta-Yahya, Mario Ouellet, Rossana Chan, Aindrila Mukhopadhyay, Jay D. Keasling, Taek Soon Lee. Identification and microbial production of a terpene-based advanced biofuel. Nature Communications, 2011; 2: 483 DOI: 10.1038/ncomms1494

Note: If no author is given, the source is cited instead.

Disclaimer: Views expressed in this article do not necessarily reflect those of ScienceDaily or its staff.

View the original article here

White matter disease: Genetic mutation causing MLC identified

ScienceDaily (May 4, 2011) — White matter disease (WMD) covers a large group of disorders that affect the white matter, or myelin. In children these disorders are commonly genetic and often go undiagnosed. In new research, a team led by Raúl Estévez, a lecturer from the Department of Physiological Sciences II, based at the UB's Bellvitge Health Sciences Campus, working with the researcher Marjo van der Knaap, from the University Medical Centre at VU University Amsterdam, have identified mutant GlialCAM as responsible for 25% of cases of megalencephalic leukoencephalopathy with subcortical cysts (MLC), a rare genetic disease affecting cerebral myelin.

Also participating in the study, which has been published and selected as a featured article in The American Journal of Human Genetics, were Tania López-Hernández, co-principal author and a postdoctoral fellow at the UB, and the researchers Albert Martínez, from the Institute of Biomedical Research (IRB Barcelona), and Virginia Nunes, a lecturer at the UB and researcher for the Bellvitge Biomedical Research Institute (IDIBELL).

Myelin is required for the correct propagation of nerve impulses between neurons, enabling the brain to send the signals that make us move. In children, diseases affecting this substance are largely genetic and affect a single gene. In adults, the diseases present as inflammatory conditions such as multiple sclerosis. "In the specific case of infant WMD, every type is rare or extremely rare, but if we consider all cases as a single group the incidence is high -- 1 patient for every 1,000 individuals," explains Raúl Estévez, ICREA Acadèmia award winner and a member of the Centre for Biomedical Network Research on Rare Diseases (CIBERER). "In addition," he adds, "in a high percentage of children with myelin disorders the diagnosis is not clear and no real conclusions can be reached."

Thanks to the identification in recent years of abnormal patterns in brain MRIs, researchers have been able to define new diseases. In 1995 experts discovered an autosomal recessive myelin disorder called megalencephalic leukoencephalopathy with subcortical cysts (MLC). In 2001 the gene responsible for 75% of the cases of this disease, MLC1, wa discovered and scientists found that other cases existed that were not caused by mutations of this gene. Of the remaining 25% of patients, two clinical phenotypes were observed: in the first case, the clinical progression, showing progressive degeneration, is the same as observed in the larger group; in the second case, the disease improves or disappears altogether. The common feature in all patients is the presence of macrocephaly, which may be accompanied by learning difficulties and autism.

The study published in The American Journal of Human Genetics takes as its starting point the genetic heterogeneity of the disease and looks for other possible mutations behind its development, combining biochemical and genetic studies. The results show that patients presenting a progressive degeneration of their condition exhibit two mutations in the GlialCAM gene, whose related protein is GlialCAM, while others exhibit only a single mutation in the same gene, which suggests a pattern of autosomal-dominant inheritance. The study, which also describes the biochemical defects observed in the disease, has revealed that mutant GlialCAM can also lead to benign familial macrocephaly and macrocephaly with mental retardation, with or without autism.

"Although we have yet to determine the exact function of GlialCAM, our research has shown that further collaborative multi-disciplinary translational studies will be required to learn more about the causes of these rare diseases and to find new treatments,"

Story Source:

The above story is reprinted (with editorial adaptations by ScienceDaily staff) from materials provided by Universidad de Barcelona, via AlphaGalileo.

Journal Reference:

Tania López-Hernández, Margreet C. Ridder, Marisol Montolio, Xavier Capdevila-Nortes, Emiel Polder, Sònia Sirisi, Anna Duarri, Uwe Schulte, Bernd Fakler, Virginia Nunes, Gert C. Scheper, Albert Martínez, Raúl Estévez, Marjo S. van der Knaap. Mutant GlialCAM Causes Megalencephalic Leukoencephalopathy with Subcortical Cysts, Benign Familial Macrocephaly, and Macrocephaly with Retardation and Autism. The American Journal of Human Genetics, 2011; 88 (4): 422 DOI: 10.1016/j.ajhg.2011.02.009

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Disclaimer: This article is not intended to provide medical advice, diagnosis or treatment. Views expressed here do not necessarily reflect those of ScienceDaily or its staff.

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Protein identified as enemy of vital tumor suppressor PTEN

ScienceDaily (May 3, 2011) — A protein known as WWP2 appears to play a key role in tumor survival, a research team headed by a scientist at The University of Texas MD Anderson Cancer Center reports in an advance online publication of Nature Cell Biology.

Their research suggests that the little-studied protein binds to the tumor-suppressing protein PTEN (phosphatase and tensin homologue deleted on chromosome 10), marking it for destruction by proteasomes, which degrade proteins and recycle their components.

PTEN plays a role regulating the cellular reproduction cycle and prevents rapid cell growth, a hallmark of malignant cells. Its gene is mutated or deleted in many types of cancer, the researchers noted.

The WWP2 (atrophin-1 interacting protein 2) protein was discovered in the laboratory of Junjie Chen, Ph.D., professor and chair in MD Anderson's Department of Experimental Radiation Oncology and senior author of the paper.

"We were trying to find regulators of PTEN when we isolated the protein WWP2 as a putative PTEN-associated protein," Chen said. He noted that WWP2 caught the researchers' attention because it is similar to the NEDD4-1 protein, which has been proposed as a regulator of PTEN function.

First suspect doesn't affect PTEN

WWP2 is an E3 ubiquitin ligase in the NEDD4-like protein family. Ubiquitins attach to other proteins, labeling them for degradation by proteasomes. NEDD4-like proteins play important roles regulating gene transcription, embryonic stem cells, cellular transport and activation of T cells.

"But when NEDD4-1 is deleted in mice, researchers have not seen a clear change in PTEN protein level," Chen noted. "These findings suggest that there may be other PTEN regulators.

"Because WWP2 is part of the NEDD4-like family, we decided to take a look at it to see if it's the real regulator of PTEN," Chen continued. "When you knock down WWP2, you see an increase in PTEN level, whereas with WWP2 overexpression you can see a decrease in PTEN. This finding indicates that WWP2 may be involved in PTEN's regulation."

Overall, the study results suggest that WWP2 can regulate PTEN stability, Chen said.

Possibly a cancer-driving gene

The team uncovered evidence that WWP2 is a potential oncogene -- a driver in tumor formation and growth. In one experiment, mice with normal WWP2 developed prostate cancer tumors after nine weeks that were more than three times the size of tumors in mice with WWP2 silenced.

Chen noted that more research is needed to determine whether WWP2 is functionally important in tumors or in tumor formation. "We need to look at real tumor samples to determine whether tumors with reduced PTEN expression could result from the overexpression of WWP2."

He added that some early studies suggest that WWP2 may operate in tumors, but a correlation between WWP2 overexpression and PTEN downregulation in tumors has not been established.

This work was supported in part by a grant from the Department of Biotechnology, Ministry of Science and Technology, India, a U.S. Department of Defense Era of Hope Research Scholar Award, an NIH Specialized Program of Research Excellence award to Mayo Clinic, and a National Cancer Institute grant to MD Anderson. Also, fellowship support came from the Department of Biotechnology, Council of Scientific and Industrial Research and University Grants Commission, India, and support from the Institute of Life Sciences, Hyderabad, India.

Co-authors with Chen are first author Subbareddy Maddika, Ph.D, Sridhar Kavela, Neelam Rani, and Vivek Reddy Palicharla, all of the Laboratory of Cell Death and Cell Survival, Centre for DNA Fingerprinting and Diagnostics in Nampally, Hyderabad, India; Jenny Pokorny and Jann Sarkaria, M.D., of the Mayo Clinic, Rochester, Minn.

Story Source:

The above story is reprinted (with editorial adaptations by ScienceDaily staff) from materials provided by University of Texas M. D. Anderson Cancer Center.

Journal Reference:

Subbareddy Maddika, Sridhar Kavela, Neelam Rani, Vivek Reddy Palicharla, Jenny L. Pokorny, Jann N. Sarkaria, Junjie Chen. WWP2 is an E3 ubiquitin ligase for PTEN. Nature Cell Biology, 2011; DOI: 10.1038/ncb2240

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Disclaimer: This article is not intended to provide medical advice, diagnosis or treatment. Views expressed here do not necessarily reflect those of ScienceDaily or its staff.

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