Neil L. Kelleher
Professor of Chemistry
Professor Kelleher received a B.S. and B.A. from Pacific Lutheran U. in 1992, a Fulbright Fellowship the following year, and a Ph.D. from Cornell U. in 1997. After a NIH Postdoctoral Fellowship at Harvard Medical School with Chris Walsh, Kelleher joined the faculty at UIUC in 1999 as a bioanalytical chemist.
Research
Research
Our laboratory has three main areas of research: custom instrumentation for Fourier Transform Mass Spectrometry (FTMS), Nuclear Signaling and Natural Products. More specifically, our main interests lie in the enzymology of natural product biosynthesis, mass spectrometric-based studies of the "Histone Code," and development of Fourier Transform Mass Spectrometry (FTMS) for Top Down Proteomics (i.e. analyzing intact proteins directly; no proteases).
A core activity is measuring chemical modifications to proteins in both hypothesis-driven and discovery modes. Our pioneering efforts in "Top Down" proteomics involve fragmenting intact protein ions in the gas phase and developing custom bioinformatics to characterize unexpected post-translational modifications (PTMs) in methane-producing microbes, yeast, and human cancer cells. In both human cell biology and antibiotic biosynthesis, key proteins harbor over 20 PTMs that present a "code" of biological logic written in the language of protein modifications. We construct, automate, and apply custom mass spectrometry and algorithms to detect and decode this logic.
Group Facilities and Instrumentation
Cell Growth
Our cell growth facility is located on the first floor of Roger Adams Lab (RAL). Here we can grow batches of cells with custom plasmids for the over-expression of enzymes involved in natural product and antibiotic synthesis.
Separations
We employ a wide range of separation methods for fractionating complex mixtures of proteins. Some of these include continuous elution gel electrophoresis (used with and acid-labile surfactant in the place of SDS), ion exchange chromatography, hydrophilic interaction chromatography (HILIC), free flow electrophoresis, and reversed phase chromatography. For nanoLC applications, we employ an Eksigent 1D system and an Advion TriVersa robot.
Mass Spectrometry
Our lab has a total of four FTMS instruments at three different locations on campus. In the main instrument bay (37 RAL), we house a custom built 8.5 Tesla Q-FTMS and a 12 Tesla LTQ-FT hybrid mass spectrometer (picture below). We also have a 7 Tesla LTQ-FT in 357 Noyes Laboratory and another 12 Tesla LTQ-FT (coming soon) in the Institute for Genomic Biology.
Bioinformatics
The new field of Top Down Proteomics has required the development of new informatic and software tools. To meet this need, we have focused thousands of hours of labor on the building and refining of the ProSight suite of software.
Publications
Mann, M., Kelleher, N.L. "Precision Proteomics: The Case for High Resolution and High Mass Accuracy" Proc. Natl. Acad. Sci. USA, 2008, epub. ahead of print: DOI 10.1073/pnas.0800788105.
Calderone, C.T., Bumpus S.B., Kelleher N.L., Walsh C.T., Magarvey N.A. "A ketoreductase domain in the PksJ protein of the bacillaene assembly line carries out both a- and ß-ketone reduction during chain growth" Proc. Natl. Acad. Sci. USA, 2008, 105, 12809–12814.
Boyne II, M.T., Garcia, B.A., Li, M., Zamdborg, L., Wenger, C.D., Babai, S., Kelleher, N.L. "Tandem Mass Spectrometry With Ultra-High Mass Accuracy Clarifies Peptide Identification by Database Retrieval" J. Proteome Res., 2008, epub. ahead of print: DOI 10.1021/pr800635m.
Wenger, C.D., Boyne II, M.T., Ferguson, J.T., Robinson, D.E., Kelleher, N.L. "A Versatile Online-Offline Engine for Automated Acquisition of High-Resolution Tandem Mass Spectra" Anal. Chem, 2008, 80, 8055-8063.
Bumpus, S.B., Kelleher, N.L. "Accessing natural product biosynthetic processes by mass spectrometry" Curr. Opin. Chem. Biol., 2008, 12, 1-8.
Bumpus, S.B., Magarvey, N.A., Kelleher, N.L., Walsh, C.T., Calderone, C.T. "Polyunsaturated Fatty-Acid-Like Trans-Enoyl Reductases Utilized in Polyketide Biosynthesis", J. Am. Chem. Soc., 2008, 130, 11614–11616.
Magarvey, N.A., Fortin, P.D., Thomas, P.M., Kelleher, N.L., Walsh, C.T. "Gatekeeping versus Promiscuity in the Early Stages of the Andrimid Biosynthetic Assembly Line", Chem. Biol., 2008, 3, 542-554.
Hatcher, N.G., Atkins Jr., N., Annangudi, S.P., Forbes, A.J., Kelleher, N.L., Gillette, M.U., Sweedler, J.V. "Mass Spectrometry-based Discovery of Circadian Peptides", Proc. Natl. Acad. Sci. USA, 2008, 105, 12527–12532.
Eliot, A.C., Griffin, B.M., Johannes, T., Thomas, P.M., Kelleher, N.L., Zhao, H., Metcalf, W.W. "Cloning, Expression, and Biochemical Characterization of Streptomyces rubellomurinus Genes Required for Biosynthesis of Antimalarial Compound FR900098", Chem. & Bio., 2008, 15, 765-770.
Brodbelt, J.S., McLafferty, F.W., Kelleher, N.L. "Focus in Honor of Roman Zubarev, Recipient of the 2007 Biemann Medal", J. Am. Soc. Mass Spectrom., 2008, 19, 751-752.
Bora, A., Annangudi, S.P., Millet, L.J., Rubakhin, S.S., Forbes, A.J., Kelleher, N.L., Gillette, M.U., Sweedler, J.V. "Neuropeptidomics of the Supraoptic Rat Nucleus", J. Proteome Res., 2008, 7, 4991-5003.
Roth, M.J., Parks, B.A., Ferguson, J.T., Li, M.T., Kelleher, N.L. "Proteotyping: Population Proteomics of Human Leukocytes Using Top Down Mass Spectrometry", Anal. Chem., 2008, 80, 2857-2866.
Garcia, B.A., Kelleher, N.L., Mizzen, C.M. "Tissue-Specific Expressions and Post-translational Modification of Histone H3 Variants", J. Proteome Res., 2008, 10, 4225-4236.
Pesavento, J.J., Bullock C.R., Leduc R.D., Mizzen, C.A., Kelleher, N.L. "Combinatorial Modification of Human Histone H4 Quantitated by Two-Dimensional Liquid Chromatography coupled with Top Down Mass Spectrometry", J. Bio. Chem., 2008, 283, 14927-14937.
Yang, H., Pesavento, J.J., Starnes, T.W., Cryderman, D.E., Wallrath, L.L., Kelleher, N.L., Mizzen, C.A. "Preferential Dimethylation of Histone H4-Lysine 20 by SUV4-20", J. Biol. Chem., 2008, 283, 12085-12092.
Zhang, J., Van Lanen, S.G., Ju, J., Liu, W., Dorrestein, P.C., Li, W., Kelleher, N.L., Shen, B. "A Phosphopantetheinylating Polyketide Synthase Producing a Linear Polyene to Initiate Enediyne Antitumor Antibiotic Biosynthesis", Proc. Natl. Acad. Sci. USA, 2008, 105, 1460-1465.
Crawford, J.M., Thomas, P.M., Scheerer, J.R., Vagstad, A.L., Kelleher, N.L., Townsend, C.A. "Deconstruction of Iterative Multidomainal Polyketide Synthase Function", Science, 2008, 320, 243-246.
Roth, M.J., Parks, B.A., Ferguson, J.T., Li, M.T., Kelleher, N.L. "Proteotyping: Population Proteomics of Human Leukocytes Using Top Down Mass Spectrometry", Anal. Chem., 2008, 80, 2857-2866.
Pesavento, J.J., Bullock C.R., Leduc R.D., Mizzen, C.A., Kelleher, N.L. "Combinatorial Modification of Human Histone H4 Quantitated by Two-Dimensional Liquid Chromatography coupled with Top Down Mass Spectrometry", J. Bio. Chem., 2008, 283, 14927-14937.
Parks, B.A., Jiang, L., Thomas, P.M., Wenger, C.D., Roth, M.J., Boyne, M.T., Burke, P.V., Kwast, K.E., Kelleher, N.L. "Top-Down Proteomics on a Chromatographic Time Scale Using Linear Ion Trap Fourier Transform Hybrid Mass Spectrometers", Anal. Chem., 2007, 79, 7984-7991.
Garcia, B.A., Pesavento, J.J., Mizzen, C.A., Kelleher, N.L. "Pervasive Combinatorial Modification of Histone H3 in Human Cells", Nat. Methods, 2007, 4, 487-489.
Hansen, D.B., Bumpus, S.B., Aron, Z.D., Kelleher, N.L., Walsh, C.T. "The Loading Module of Mycosubtilin: An Adenylation Domain with Fatty Acid Selectivity." J. Am. Chem. Soc., 2007, 129, 6366-6367
Jiang, L., Smith, J.N., Mizzen, C.A., Kelleher, N.L. "Global Assessment of Combinatorial Post-translational Modification of Core Histones in Yeast Using Contemporary Mass Spectrometry", J. Biol. Chem., 2007, 282, 27923-27934.
Awards
- University Scholar, 2009-2010
- Biemann Medal, 2009
- Pittsburgh Conference Achievement Award, 2008
- Pfizer Award in Enzyme Chemistry from the ACS Division of Biological Chemistry, 2007
- Presidential Early Career Award
- Alfred P. Sloan Fellow
- Packard Fellow
- NSF CAREER Award
- Lilly Analytical Chemistry Award
- Burroughs Wellcome Fund Young Investigator
- Searle Scholar
- Fulbright Scholar
Highlights
1. First-of-a-kind Mass Spectrometers now up and running.
Both 12 Tesla LTQ FT mass spectrometers are now on-line and producing prodigious amounts of data for both small molecules (natural product discovery) and proteins (studying human cancer cells as they enter anti-proliferative states of growth arrest and controlled cell death).
2. New Paper in Science Magazine.
Together with the Townsend Group at Johns Hopkins, we published an article in Science on the biosynthesis of aflatoxin. Here the function of a new enzyme domain was determined and direct observation of intermediates from an "iterative" polyketide synthase was accomplished using high performance mass spectrometry.
3. "Top Down" Mass Spec. goes high throughput.
In a series of two articles in Analytical Chemistry, we have demonstrated the capability to accomplish large scale analysis of intact proteins from yeast cells in culture and human white blood cells. HIGH-RESOLUTION tandem mass spectrometry of intact proteins is now feasible on a chromatographic time scale, with all the associated software ready for even larger scale Top Down Proteome Projects.
- Read the Analytical Chemistry article.
- A review of this material was done in the Journal of Proteome Research, Vol. 7, No. 5, 2008.
