Techology development

Our co-location with the Joint Bioenergy Institute gives our lab access to state-of-the-art mass spectrometry and robotics facilities. We develop novel technologies and apply them towards engineering microbes.

  • We are developing mass spectrometry-based tools for measuring metabolite and protein abundance, and using these tools to “debug” our metabolic pathways.
  • Likewise we are developing methods for the prediction of production phenotypes using metabolic flux, proteomic, and transcriptomic data gathered by our in-house facilities.
  • In collaboration with the JBEI proteomics group, we have developed robust LC tandem MS methods to monitor enzyme-tethered intermediates in polyketide synthases.
  • We are actively involved in high-throughput analytical chemistry method development and applying these methods to further our enzyme discovery and metabolic engineering efforts.
  • Finally, we are developing tools for rapid de novo DNA synthesis to enable faster strain engineering.

FUNDING

National Science Foundation, DARPA, Department of Energy

REPRESENTATIVE PUBLICATIONS
  • M. M. Maharbiz, W. J. Holtz, R. T. Howe, and J. D. Keasling. 2004. “Microbioreactor arrays with parametric control for high-throughput experimentation.” Biotechnol. Bioeng. 86:485-90.
  • A. S. Shaikh, Y, J. Tang, A. Mukhopadhyay, and J. D. Keasling. 2008. “Isotopomer distributions in amino acids from a high expressed protein as a proxy for those from total protein.” Anal. Chem. 80:886-890.
  • E. E. K. Baidoo, P. I. Benke, C. Neususs, M. Pelzing, G. Kruppa, J. A. Leary, and J. D. Keasling. 2008. “Capillary electrophoresis-Fourier transform ion cyclotron resonance mass spectrometry for the identification of cationic metabolites via a pH-mediated stack-transient isotrachophoretic method.” Anal. Chem. 80:3112-3122.
  • D. E. Garcia, E. E. Baidoo, P. I. Benke, F. Pingitore, Y. J. Tang, S. Villa, and J. D. Keasling. 2008. “Separation and mass spectrometry in microbial metabolomics.” Curr. Opin. Microbiol. 11:233-239.
  • S. K. Lee and J. D. Keasling. 2008. “Heterologous protein production in Escherichia coli using the propionate-inducible pPro system by conventional and auto-induction methods.” Prot. Exp. Purif. 61:197-203.
  • Y. J. Tang, H. Garcia Martin, S. Myers, S. Rodriguez, E. E. K. Baidoo, and J. D. Keasling. 2009. “Advances in analysis of microbial metabolic fluxes via 13C isotopic labeling.” Mass Spec. Rev. 28:362-375.
  • W. D. Marner, A. Shaikh, S. J. Muller, and J. D. Keasling. 2009. “Enzyme immobilization via silaffin-mediated autoencapsulation in a biosilica support.” Biotech. Prog. 25:417-423.
  • M. Ouellet, P. D. Adams, J. D. Keasling, and A. Mukhopadhyay. 2010. “A rapid and inexpensive method for microarray gene expression analysis.” BMC Biotechnol. 9:97 (doi:10.1186/1472-6750-9-97).
  • Poust, S., Phelan, R.M., Deng, K., Katz, L., Petzold, C.J. and Keasling, J.D., 2015. Divergent Mechanistic Routes for the Formation of gem‐Dimethyl Groups in the Biosynthesis of Complex Polyketides. Angewandte Chemie International Edition, 54(8), pp.2370-2373.
  • A. S. Shaikh, Y. J. Tang, A. Mukhopadhyay, H. G. Martín, J. Gin, P. Benke, and J. D. Keasling. 2010. “Study of stationary phase metabolism via isotopomer analysis of amino acids from an isolated protein.” Biotechnol. Prog. 26:52-56.
  • A. M. Redding-Johanson, T. S. Batth, R. Chan, R. Krupa, H. L. Szmidt, P. D. Adams, J. D. Keasling, T. S. Lee, A. Mukhopadhyay, C. J. Petzold. 2011. “Targeted proteomics for metabolic pathway optimization: application to terpene production.” Metab. Eng. 13:194-203.
  • T. S. Lee, R. A. Krupa, F. Zhang, M. Hajimorad, W. J. Holtz, N. Prasad, S. K. Lee, and J. D. Keasling. 2011. “BglBrick vectors and datasheets: a synthetic biology platform for gene expression.” J. Biol. Eng. 5:12.
  • S. R. Chhabra, G. Butland, D. Elias, J.-M. Chandonia, O.-Y. Fok, T. Juba, A. Gorur, S. Allen, C. M. Leung, K. Keller, S. Reveco, G. Zane, E. Semkiw, R. Prathapam, B. Gold, M. Singer, M. Ouellet, E. Szakal, D. Jorgens, M. Price, E. Witkowska, H. Beller, T. C. Hazen, M. D. Biggin, M. Auer, J. Wall, and J. D. Keasling. 2011. “Generalized schemes for high throughput manipulation of the Desulfovibrio vulgaris Hildenborough genome”. Appl. Environ. Microbiol. 77:7595-7604.
  • T. de Rond, P. Peralta-Yahya, X. Cheng, T. R. Northen, and J. D. Keasling. 2013. “Versatile synthesis of probes for high throughput enzyme activity screening.” Anal. Bioanal. Chem. 405:4969-4973.
  • T. S. Batth, P. Singh, V. R. Ramakrishnan, M. M. Sousa, L. J. Chan, H. M Tran, E. G. Luning, E. H. Pan, K. M. Vuu, J. D. Keasling, P. D. Adams, and C. J. Petzold. 2014. “A targeted proteomics toolkit for high-throughput absolute quantification of Escherichia coli proteins.” Met. Eng. 26:48-59.
  • Poust, S., Phelan, R.M., Deng, K., Katz, L., Petzold, C.J. and Keasling, J.D., 2015. Divergent Mechanistic Routes for the Formation of gem‐Dimethyl Groups in the Biosynthesis of Complex Polyketides. Angewandte Chemie International Edition, 54(8), pp.2370-2373.
  • Hagen, A., Poust, S., Rond, T.D., Fortman, J.L., Katz, L., Petzold, C.J. and Keasling, J.D., 2016. Engineering a polyketide synthase for in vitro production of adipic acid. ACS synthetic biology.