Organism development and characterization for ethanol production using thermophilic bacteria

  • Taryn R. Klapatch
  • David A. L. Hogsett
  • Sunitha Baskaran
  • Sunita Pal
  • Lee R. Lynd
Session 2 Applied Biological Research

Abstract

This article provides an overview and evaluation of our recent interdisciplinary work having a bearing on ethanol production using thermophilic bacteria. Based on steady-state effluent substrate concentrations in continuous culture, the ratio of the ethanol-inhibited growth rate and the uninhibited growth rate (ΜI0)s was determined forClostridium thermosaccharolyticum grown on xylose at both 55 and 60 ‡C. (ΜI0)S exhibited an approximately linear pattern in relation to ethanol concentration. Based on least square linear fits to the data, ethanol concentrations corresponding to (ΜI0S = 0.5 were 29 g/L ethanol at 60‡C, and 36 g/L ethanol at 55‡C, and 31 g/L for the data combined for both temperatures. It is concluded that ethanol inhibition is unlikely to constrain utilization ofC. thermosaccharolyticum in processes for ethanol production from cellulosic biomass. Growth at high substrate concentrations using a defined medium has been achieved forC. thermocellum in continuous culture with essentially complete substrate utilization at 54 g/L cellobiose concentration. We also include work aimed at elucidating the molecular genetics ofC. thermocellum with the ultimate goal of pathway manipulation. Antibiotics effective againstC. thermocellum include chloramphenicol, thiamphenicol, and erythromycin at 125, 10, and 40 Μg/mL, respectively. Summarized is work describing a restriction system ofC. thermocellum and protection against it by methylation. Demonstration of restriction protection and antibiotic sensitivity provides two elements likely to be useful in the transformation ofC. thermocellum.

Index Entries

Clostridium thermocellum Clostridium thermosac-charolyticum ethanol production 

References

  1. 1.
    Johnson, E. A., Sakojoh, M., Hallivvell, G., Madia, A., and Dermain, A. L. (1982),Appl. Env. Microbiol. 43(5), 1125.Google Scholar
  2. 2.
    Lamed R. and Bayer, E. A. (1988),Adv. Appl. Microbiol. 33, 1.CrossRefGoogle Scholar
  3. 3.
    Lynd, L. R., Wolkin, R. H., and Grethlein, H. E. (1989),Appl. Env. Microbiol. 55, 3131.Google Scholar
  4. 4.
    Ahn, A. J. (1991), M.S. Thesis, Thayer School of Engineering, Dartmouth College, Hanover, NH.Google Scholar
  5. 5.
    Lynd, L. R. (1989),Adv. Biochem. Engr. Biotechnol. 38, 1.Google Scholar
  6. 6.
    Hogsett, D. A., Ahn, H.-J., South, C. R., and Lynd, L. R. (1992),Appl. Biochem. Biotechnol. 34/35, 527.CrossRefGoogle Scholar
  7. 7.
    Stone, K. and Lynd, L. R. (1993), to appear inProceedings of the Annual Automotive Technology Contractors Coordination Meeting. Society of Automotive Engineers.Google Scholar
  8. 8.
    Lynd, L. R., Ahn, H.-J., Anderson, G., Hill, P., Kersey, D. S., Klapatch, T. A. (1991),Appl. Biochem. Biotechnol. 28/29, 549.CrossRefGoogle Scholar
  9. 9.
    Klapatch, T. R. and Lynd, L. R. (1993), in preparation.Google Scholar
  10. 10.
    Beaven, M. J., Charpentier, C., and Rose, A. H. (1982),J. Gen. Microbiol. 128, 1447.Google Scholar
  11. 11.
    Loureiro, V. and Ferreira, H. G. (1983),Biotechnol. Bioeng. 25, 2263.CrossRefGoogle Scholar
  12. 12.
    Nagodawithana, T. W. and Steinkraus, K. H. (1976),Appl. Env. Microbiol. 31, 158.Google Scholar
  13. 13.
    Novak, M., Strehaiano, P., Moreno, M., and Goma, G. (1981),Biotechnol. Bioeng. 23, 201.CrossRefGoogle Scholar
  14. 14.
    Dasari, G., Roddick, F., Connor, M. A., and Pamment, N. B. (1983),Biotechnol. Lett. 5, 715.CrossRefGoogle Scholar
  15. 15.
    Dombek, K. M. and Ingram, L. O. (1986),Appl. Env. Microbiol. 51, 197.Google Scholar
  16. 16.
    Guijarro, J. M. G., and Lapunas, R. (1984),J. Bacteriol. 160, 874.Google Scholar
  17. 17.
    Jobses, I. M. L. and Roels, J. A. (1986),Biotechnol. Bioeng. 28, 554.CrossRefGoogle Scholar
  18. 18.
    Casey, G. P., Magnus, C. A., and Ingledew, W. M. (1983),Biotechnol. Lett. 5, 249.CrossRefGoogle Scholar
  19. 19.
    D’Amore, T., Panchal, C. J., Russell, I., and Stewart, G. G. (1988),J. Ind. Microbiol. 2, 365.CrossRefGoogle Scholar
  20. 20.
    Dasari, G., Worth, M. A., Connor, M. A., and Pamment, N. B. (1990),Biotechnol. Bioeng. 35, 109.CrossRefGoogle Scholar
  21. 22.
    Aiba, S., Shoda, M., and Nagatani, M. (1968),Biotechnol. Bioeng. 10, 845.CrossRefGoogle Scholar
  22. 22.
    Bazua, C. D. and Wilke, C. R. (1967),Biotechnol. Bioeng. Symp. 7, 105.Google Scholar
  23. 23.
    Ghose, T. K. and Tyagi, R.D. (1979),Biotechnol. Bioeng. 21, 1401.CrossRefGoogle Scholar
  24. 24.
    Holzberg, I., Finn, R. K., and Steinkraus, K. H. (1967),Biotechnol. Bioeng. 9, 413.CrossRefGoogle Scholar
  25. 25.
    Lee, K. J. and Rogers, P. L. (1983),Chem. Eng. J. 27, B31.CrossRefGoogle Scholar
  26. 26.
    Levenspiel, O. (1980),Biotechnol. Bioeng. 22, 671.CrossRefGoogle Scholar
  27. 27.
    Worden, R. M., Donalson, T. L., and Shumate, S. E. (1983),Biotechnol. Bioeng. Symp. 13, 265.Google Scholar
  28. 28.
    Benschoter, A. S. and Ingram, L. O. (1986),Appl. Environ. Microbiol. 51, 1278.Google Scholar
  29. 29.
    Herrero, A. A. and Gomez, R. F. (1980),Appl. Env. Microbiol. 40(3), 571.Google Scholar
  30. 30.
    Lovitt, R. W., Longin, R., and Zeikus, J. G. (1984),Appl. Environ. Microbiol. 48, 171.Google Scholar
  31. 31.
    Sinensky, M. (197),Proc. Natl. Acad. Sci. USA 71, 522.Google Scholar
  32. 32.
    van Uden, N. (1985),Annual Reports on Fermentation Processes 8, 11.Google Scholar
  33. 33.
    van Uden, N. (1984),Adv. Microb. Phys. 25, 195.Google Scholar
  34. 34.
    Huang, S.-Y. and Chen, J.-C. (1988),Enz. Microb. Technol. 20, 431.CrossRefGoogle Scholar
  35. 35.
    Chem Systems (1992),Technical & Economic Evaluation, Wood to Ethanol Process, Office of Energy Demand Policy, DOE, In Press.Google Scholar
  36. 36.
    South, C. R. and Lynd, L. R. (1993),Appl. Biochem. Biotechnol., this volume.Google Scholar
  37. 37.
    Lynd, L. R., Cushman, J., Nichols, R., and Wyman, C. E. (1991),Science 251, 1318.CrossRefGoogle Scholar
  38. 38.
    Hill, P. W., Klapatch, T. R., and Lynd, L. R. (1993),Biotechnol. Bioeng. 42(7), 873.CrossRefGoogle Scholar
  39. 39.
    Wang, D. I. C. and Dalal, R. (1986), US Patent No. 4,568,644.Google Scholar
  40. 40.
    Vancanneyt, M., De Vos, P., and De Ley, J. (1987),J. Biotechnol. Lett 9(8), 567.CrossRefGoogle Scholar
  41. 41.
    Liu, H.-S., Hsu, H.-W., and Sayler, G. (1988),Biotechnol. Prog. 4, 40.CrossRefGoogle Scholar
  42. 42.
    Mistry, F. and Cooney, C. L. (1989),Biotechnol. Bioeng. 34, 1295.CrossRefGoogle Scholar
  43. 43.
    Wiegel, J., Ljungdahl, L. G., and Rawson, J. R. (1979),J. Bacterial. 139, 800.Google Scholar
  44. 44.
    Ben-Bassat, A., Lamed, R., Ng, T. K., and Zeikus, J. G. (1980),Energy from Biomass and Wastes IV, Institute of Gas Technology, Chicago, p. 275.Google Scholar
  45. 45.
    Hyun, H. H., Shen, G.-J., and Zeikus, J. G. (1985),J. Bacteriol. 164, 1153.Google Scholar
  46. 46.
    Sundquist, J. A., Blanch, H. W., and Wilke, C. R. (1986), Presented at the 192nd meeting of the ACS.Google Scholar
  47. 47.
    Toukourou, F., Donaduzzi, L., Miclo, A., and Germain, P. (1989),Biotechnol. Lett. 11, 443.CrossRefGoogle Scholar
  48. 48.
    Mori, Y. and Inaba, T. (1990),Biotechnol. Bioeng. 36, 849.CrossRefGoogle Scholar
  49. 49.
    Lamed, R. and Zeikus, J. G. (1980),J. Bacteriol. 144, 569.Google Scholar
  50. 50.
    Wang, D. I. C., Avgerinos, G. C., Biocic, I., Fang, S. D., and Fang, H. Y. (1983),Philos. Trans. R. Soc. Lond. B300, 323.CrossRefGoogle Scholar
  51. 51.
    Brener, D. and Johnson, B. F. (1984),Appl. Environ. Microbiol. 47, 1126.Google Scholar
  52. 52.
    Bender, J., Vatcharapijarn, Y., and Jeffries, T. W. (1985),Appl. Environ. Microbiol. 49, 475.Google Scholar
  53. 53.
    Kurose, N., Yagyu, J., Miyazaki, T., Uchida, M., Hanai, S., and Obayashi, A. (1986),J. Ferment. Technol. 64, 447.CrossRefGoogle Scholar
  54. 54.
    Venkateswaran, S. and Demain, A. L. (1986),Chem. Eng. Commun. 45, 53.CrossRefGoogle Scholar
  55. 55.
    Hormeyer, H. F., Tailliez, P., Millet, J., Girard, H., Bonn, G., Bobleter, O., and Aubert, J.-P. (1988),Appl. Microbiol. Biotechnol. 29, 528.CrossRefGoogle Scholar
  56. 56.
    Lamed, R., Lobos, J. H., and Su, T.M. (1988),Appl. Environ. Microbiol. 54, 1216.Google Scholar
  57. 57.
    Hon-Nami, K., Goto, S., Tomita, M., Takagi, Y., Sekine, K., Okuma, E., Yonemura, S., Sato, K., and Saiki, T. (1988),Proceedings of the 8th International Symposium on Alcohol Fuels. 71–76.Google Scholar
  58. 58.
    Sai-Ram, M., Seenayya, G. (1989),Biotechnol. Lett. 11, 589.CrossRefGoogle Scholar
  59. 59.
    Tailliez, P., Girard, H., Longin, R., Beguin, P., and Millet, J. (1989),Appl. Environ. Microbiol. 55, 203.Google Scholar
  60. 60.
    Ljungdahl, L. G., Bryant, F., Carreira, L., Saiki, T., and Wiegel, J. (1981),Trends in the Biology of Fermentations for Fuels and Chemicals. Hollaender, A., Rabson, R., Rogers, P., San Pietro, A., Valentine, R., and Wolfe, R. (eds.), Plenum, New York, p. 397.Google Scholar
  61. 61.
    Camera, L. H., Wiegel, J., and Ljungdahl, L. G. (1983),Biotechnol. Bioeng. Symp. 13, 183.Google Scholar
  62. 62.
    Ljungdahl, L. G., and Camera, L. H. (1983), US Patent No. 4,385,117.Google Scholar
  63. 63.
    Kannan, V. and Mutharasan, R. (1985),Enz. Microb. Technol. 7, 87.CrossRefGoogle Scholar
  64. 64.
    Lacis, L. S. and Lawford, H. G. (1988),Arch. Microbiol. 150, 48.CrossRefGoogle Scholar
  65. 65.
    Lacis, L. S. and Lawford, H. G. (1991),Appl. Environ. Microbiol. 57, 579.Google Scholar
  66. 66.
    Rogers, P. and Gottschalk. G. (1993),Biochemistry and Regulation of Acid and Solvent Production in Clostridia. Chapter 2, in press.Google Scholar
  67. 67.
    Mermelstein, L. D., Welker, N. E., Bennett, G. N., and Papoutsakis, E. T. (1992),Bio/Technol. 10, 190.CrossRefGoogle Scholar
  68. 68.
    Mermelstein, L. D. and Papoutsakis, E. T. (1993),Appl. Environ. Microbiol. 59, 1077.Google Scholar
  69. 69.
    Young, M., Minton, N. P., and Staudenbauer, W. L. (1989),FEMS Microbiol. Rev. 63, 301.CrossRefGoogle Scholar
  70. 70.
    Peteranderl, R., Shotts, Jr., E. B., and Wiegel, J. (1990),Appl. Environ. Microbiol. 56, 1981.Google Scholar
  71. 71.
    Avgerinos, G. C. (1982), Ph.D. Thesis, MIT.Google Scholar

Copyright information

© Humana Press Inc. 1994

Authors and Affiliations

  • Taryn R. Klapatch
    • 1
  • David A. L. Hogsett
    • 2
  • Sunitha Baskaran
    • 2
  • Sunita Pal
    • 3
  • Lee R. Lynd
    • 1
    • 2
  1. 1.Department of Biological SciencesDartmouth CollegeHanover
  2. 2.Thayer School of EngineeringDartmouth CollegeHanover
  3. 3.Department of BiochemistryDartmouth CollegeHanover

Personalised recommendations