An Engineered Microbial Platform for Direct Biofuel Production from Brown Macroalgae
Sourced from Seaweed
Using seaweed as a raw material for biofuels has received relatively little attention, in part because their primary sugar constituent, alginate, is not readily fermented by industrially tractable microbes. Wargacki et al. (p. 308; see the cover) now demonstrate that metabolically engineered bacteria can degrade seaweed and subsequently ferment the sugars into ethanol at laboratory scale.
Abstract
Prospecting macroalgae (seaweeds) as feedstocks for bioconversion into biofuels and commodity chemical compounds is limited primarily by the availability of tractable microorganisms that can metabolize alginate polysaccharides. Here, we present the discovery of a 36–kilo–base pair DNA fragment from Vibrio splendidus encoding enzymes for alginate transport and metabolism. The genomic integration of this ensemble, together with an engineered system for extracellular alginate depolymerization, generated a microbial platform that can simultaneously degrade, uptake, and metabolize alginate. When further engineered for ethanol synthesis, this platform enables bioethanol production directly from macroalgae via a consolidated process, achieving a titer of 4.7% volume/volume and a yield of 0.281 weight ethanol/weight dry macroalgae (equivalent to ~80% of the maximum theoretical yield from the sugar composition in macroalgae).
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References and Notes
1
Stephanopoulos G., Challenges in engineering microbes for biofuels production. Science 315, 801 (2007).
2
G. Roesijadi, S. B. Jones, L. J. Snowden-Swan, Y. Zhu, “Macroalgae as a Biomass Feedstock: A Preliminary Analysis,” prepared for the U.S. Department of Energy under contract DE-AC05-76RL01830 by Pacific Northwest National Laboratory (2010).
3
Somerville C., Youngs H., Taylor C., Davis S. C., Long S. P., Feedstocks for lignocellulosic biofuels. Science 329, 790 (2010).
4
Adams J. M., Gallagher J., Donnison I. S., Fermentation study on Saccharina latissima for bioethanol production considering variable pre-treatments. J. Appl. Phycol. 21, 569 (2008).
5
Horn I. M. A. S. J., Ostgaard K., Production of ethanol from mannitol by Zymobacter palmae. J. Ind. Microbiol. Biotechnol. 24, 51 (2000).
6
Horn S. J., Aasen I. M., Ostgaard K., Ethanol production from seaweed extract. J. Ind. Microbiol. Biotechnol. 25, 249 (2000).
7
K. I. Draget, O. Smidsrod, G. Skjak-Braek, in Polysaccharides and Polyamides in the Food Industry. Properties, Production, and Patents, A. Steinbuchel, S. K. Rhee, Eds. (Wiley–VCH Verlag GmbH KGaA, Weiheim, 2005), pp. 1–30.
8
Wong T. Y., Preston L. A., Schiller N. L., Alginate lyase: Review of major sources and enzyme characteristics, structure-function analysis, biological roles, and applications. Annu. Rev. Microbiol. 54, 289 (2000).
9
Ochiai A., Yamasaki M., Mikami B., Hashimoto W., Murata K., Crystallization and preliminary x-ray analysis of an exotype alginate lyase Atu3025 from Agrobacterium tumefaciens strain C58, a member of polysaccharide lyase family 15. Acta Crystallogr. Sect. F Struct. Biol. Cryst. Commun. 62, 486 (2006).
10
Preiss J., Ashwell G., Alginic acid metabolism in bacteria. II. The enzymatic reduction of 4-deoxy-L-erythro-5-hexoseulose uronic acid to 2-keto-3-deoxy-D-gluconic acid. J. Biol. Chem. 237, 317 (1962).
11
Takase R., Ochiai A., Mikami B., Hashimoto W., Murata K., Molecular identification of unsaturated uronate reductase prerequisite for alginate metabolism in Sphingomonas sp. A1. Biochim. Biophys. Acta 1804, 1925 (2010).
12
Takeda H., Yoneyama F., Kawai S., Hashimoto W., Murata K., Bioethanol production from marine biomass alginate by metabolically engineered bacteria. Energy Environ. Sci. 4, 2575 (2011).
13
Alper H., Stephanopoulos G., Engineering for biofuels: Exploiting innate microbial capacity or importing biosynthetic potential? Nat. Rev. Microbiol. 7, 715 (2009).
14
Atsumi S., Hanai T., Liao J. C., Non-fermentative pathways for synthesis of branched-chain higher alcohols as biofuels. Nature 451, 86 (2008).
15
Bond-Watts B. B., Bellerose R. J., Chang M. C., Enzyme mechanism as a kinetic control element for designing synthetic biofuel pathways. Nat. Chem. Biol. 7, 222 (2011).
16
Dellomonaco C., Clomburg J. M., Miller E. N., Gonzalez R., Engineered reversal of the β-oxidation cycle for the synthesis of fuels and chemicals. Nature 476, 355 (2011).
17
Ohta K., Beall D. S., Mejia J. P., Shanmugam K. T., Ingram L. O., Genetic improvement of Escherichia coli for ethanol production: Chromosomal integration of Zymomonas mobilis genes encoding pyruvate decarboxylase and alcohol dehydrogenase II. Appl. Environ. Microbiol. 57, 893 (1991).
18
Lee H. Y., Harvey C. J., Cane D. E., Khosla C., Improved precursor-directed biosynthesis in E. coli via directed evolution. J. Antibiot. (Tokyo) 64, 59 (2011).
19
Pfeifer B. A., Admiraal S. J., Gramajo H., Cane D. E., Khosla C., Biosynthesis of complex polyketides in a metabolically engineered strain of E. coli. Science 291, 1790 (2001).
20
Ajikumar P. K., et al., Isoprenoid pathway optimization for Taxol precursor overproduction in Escherichia coli. Science 330, 70 (2010).
21
Alper H., Miyaoku K., Stephanopoulos G., Construction of lycopene-overproducing E. coli strains by combining systematic and combinatorial gene knockout targets. Nat. Biotechnol. 23, 612 (2005).
22
Leonard E., et al., Strain improvement of recombinant Escherichia coli for efficient production of plant flavonoids. Mol. Pharm. 5, 257 (2008).
23
Martin V. J., Pitera D. J., Withers S. T., Newman J. D., Keasling J. D., Engineering a mevalonate pathway in Escherichia coli for production of terpenoids. Nat. Biotechnol. 21, 796 (2003).
24
Steen E. J., et al., Microbial production of fatty-acid–derived fuels and chemicals from plant biomass. Nature 463, 559 (2010).
25
Schirmer A., Rude M. A., Li X., Popova E., del Cardayre S. B., Microbial biosynthesis of alkanes. Science 329, 559 (2010).
26
Skraly F. A., Lytle B. L., Cameron D. C., Construction and characterization of a 1,3-propanediol operon. Appl. Environ. Microbiol. 64, 98 (1998).
27
Yim H., et al., Metabolic engineering of Escherichia coli for direct production of 1,4-butanediol. Nat. Chem. Biol. 7, 445 (2011).
28
Lynd L. R., van Zyl W. H., McBride J. E., Laser M., Consolidated bioprocessing of cellulosic biomass: An update. Curr. Opin. Biotechnol. 16, 577 (2005).
29
Li J. W., et al., Purification and characterization of a bifunctional alginate lyase from Pseudoalteromonas sp. SM0524. Mar. Drugs 9, 109 (2011).
30
He S. Y., Lindeberg M., Chatterjee A. K., Collmer A., Cloned Erwinia chrysanthemi out genes enable Escherichia coli to selectively secrete a diverse family of heterologous proteins to its milieu. Proc. Natl. Acad. Sci. U.S.A. 88, 1079 (1991).
31
Kazemi-Pour N., Condemine G., Hugouvieux-Cotte-Pattat N., The secretome of the plant pathogenic bacterium Erwinia chrysanthemi. Proteomics 4, 3177 (2004).
32
Fujiyama K., Maki H., Kinoshita S., Yoshida T., Purification and characterization of the recombinant alginate lyase from Pseudomonas sp. leaked by Escherichia coli upon addition of glycine. FEMS Microbiol. Lett. 126, 19 (1995).
33
Klemm P., Hjerrild L., Gjermansen M., Schembri M. A., Structure-function analysis of the self-recognizing Antigen 43 autotransporter protein from Escherichia coli. Mol. Microbiol. 51, 283 (2004).
34
van der Woude M. W., Henderson I. R., Regulation and function of Ag43 (flu). Annu. Rev. Microbiol. 62, 153 (2008).
35
Henderson I. R., Owen P., The major phase-variable outer membrane protein of Escherichia coli structurally resembles the immunoglobulin A1 protease class of exported protein and is regulated by a novel mechanism involving Dam and oxyR. J. Bacteriol. 181, 2132 (1999).
36
Kjaergaard K., Hasman H., Schembri M. A., Klemm P., Antigen 43-mediated autotransporter display, a versatile bacterial cell surface presentation system. J. Bacteriol. 184, 4197 (2002).
37
Hashimoto W., Kawai S., Murata K., Bacterial supersystem for alginate import/metabolism and its environmental and bioenergy applications. Bioeng. Bugs 1, 97 (2010).
38
Hugouvieux-Cotte-Pattat N., Reverchon S., Two transporters, TogT and TogMNAB, are responsible for oligogalacturonide uptake in Erwinia chrysanthemi 3937. Mol. Microbiol. 41, 1125 (2001).
39
Hashimoto W., Miyake O., Momma K., Kawai S., Murata K., Molecular identification of oligoalginate lyase of Sphingomonas sp. strain A1 as one of the enzymes required for complete depolymerization of alginate. J. Bacteriol. 182, 4572 (2000).
40
Ochiai A., Hashimoto W., Murata K., A biosystem for alginate metabolism in Agrobacterium tumefaciens strain C58: Molecular identification of Atu3025 as an exotype family PL-15 alginate lyase. Res. Microbiol. 157, 642 (2006).
41
Blot N., Berrier C., Hugouvieux-Cotte-Pattat N., Ghazi A., Condemine G., The oligogalacturonate-specific porin KdgM of Erwinia chrysanthemi belongs to a new porin family. J. Biol. Chem. 277, 7936 (2002).
42
Lau M. W., Dale B. E., Cellulosic ethanol production from AFEX-treated corn stover using Saccharomyces cerevisiae 424A(LNH-ST). Proc. Natl. Acad. Sci. U.S.A. 106, 1368 (2009).
43
Yomano L. P., York S. W., Shanmugam K. T., Ingram L. O., Deletion of methylglyoxal synthase gene (mgsA) increased sugar co-metabolism in ethanol-producing Escherichia coli. Biotechnol. Lett. 31, 1389 (2009).
44
Datsenko K. A., Wanner B. L., One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products. Proc. Natl. Acad. Sci. U.S.A. 97, 6640 (2000).
45
Zhang Z., et al., Preparation and structure elucidation of alginate oligosaccharides degraded by alginate lyase from Vibro sp. 510. Carbohydr. Res. 339, 1475 (2004).
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Published In
Science
Volume 335 | Issue 6066
20 January 2012
20 January 2012
Copyright
Copyright © 2012, American Association for the Advancement of Science.
Submission history
Received: 27 September 2011
Accepted: 15 November 2011
Published in print: 20 January 2012
Acknowledgments
This work is supported by the DOE under Advanced Research Projects Agency–Energy (ARPA-E) award DE-AR0000006 and by the CORFO INNOVA CHILE (código 09CTEI-6866). We thank M. Polz (Department of Civil and Environmental Engineering, Massachussetts Institute of Technology) for the kind gift of the V. splendidus strain. We thank R. Bailey for important discussion on microbial engineering. We also thank A. Wahler for critical review and editing of the manuscript. The assistance of A. Gill for in vitro characterization of Oal enzymes is greatly appreciated. Patents describing components of this work can be found under U.S. patent application nos. 12/245537, 12/361293, 12/899419, 61/427077, and 61/436173. This report was prepared as an account of work sponsored by an agency of the U.S. government. Neither the U.S. government nor any agency thereof, nor any of their employees, makes any warranty, express or implied, or assumes any legal liability or responsibility for the accuracy, completeness, or usefulness of any information, apparatus, product, or process disclosed, or represents that its use would not infringe privately owned rights. Reference herein to any specific commercial product, process, or service by trade name, trademark, manufacturer, or otherwise does not necessarily constitute or imply its endorsement, recommendation, or favoring by the U.S. government or any agency thereof. The views and opinions of authors expressed herein do not necessarily state or reflect those of the U.S. government or any agency thereof.
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