Charles W. Gehrke Proteomics Center
E-mail: mooneyb at missouri dot edu
Web site: Charles W. Gehrke Proteomics Center
Office address: 214 Bond Life Sciences Center
Office phone: 573-884-7374
The polyhydroxyalkanoate (PHA) copolymer, P(3HB-co-3HV), is a type of biodegradable plastic demonstrated to be useful in the manufacture of bottles, bags, disposable razors, flatware, etc. Additionally, medical uses for PHA, e.g. implants, drug delivery, have been proposed. To date there has been limited success in using plants as a means of PHA production by targeting the bacterial enzymes responsible for PHA synthesis to the chloroplasts of Arabidopsis thaliana. This resulted in the production of low levels (3% dry wt.) of the PHA copolymer in the chloroplasts. One of the precursors for the PHA co-polymer, acetyl-CoA, is present naturally in the chloroplasts at sufficient levels to support PHA production. However, the other precursor, propionyl-CoA, was produced at a very low efficiency by the endogenous plastid pyruvate dehydrogenase complex (PDC). The branched-chain α-ketoacid dehydrogenase enzyme (BCE1) is capable of production of propionyl-CoA at a much higher efficiency that the plastid PDC. However the BCE1 enzyme is located in a different compartment of the plant cell, the mitochondria.
The objective of the project (being conducted in Douglas D. Randall's lab by Elizabeth Hoyos) is to alter the intra-cellular location of the Arabidopsis thaliana BCE1 (AtBCE1) enzyme from mitochondria to plastids. We have generated plastid-targeted AtBCE1 by replacing the mitochondrial-targeting portions of the two proteins that make up the AtBCE1 (AtBCE1α and AtBCE1β) with plastid-targeting sequences derived from the A. thaliana small subunit of RuBisCO. In addition, we have replaced the region of AtBCE1βresponsible for binding to the central core enzyme, BCE2, with a region capable of binding plastid PDC E2. We have confirmed plastid-targeting by western blot of chloroplasts isolated from transgenic plants. We have also confirmed (although indirectly through enzyme assays) the ability of the plastid-targeted AtBCE1 to form a heterotetrameric complex as well as its ability to bind to the endogenous plastid PDC E2. Once we have confirmed stable, heritable, and functional, plastid-targeted AtBCE1 we will cross these plants with those that posses the (bacterial) genes for biodegradable plastic synthesis. We will then asses levels of biodegradable plastic produced in these transgenic plants.
Yao H, Kato A, Mooney B, Birchler JA. Phenotypic and gene expression analyses of a ploidy series of maize inbred Oh43. Plant Molecular Biology 2010 (in press, DOI: 10.1007/s11103-010-9722-4).
Sauter ER, Davis W, Qin W, Scanlon S, Mooney B, Bromert K and Folk WR. Identification of a beta-casein-like peptide in breast nipple aspirate fluid that is associated with breast cancer. Biomarkers in Medicine 2009;3(5):577-588.
Mooney BP. The second green revolution? Production of plant-based biodegradable plastics. Biochemical Journal 2009;418(2):219-232.
Chen M, Mooney BP, Hajduch M, Joshi T, Zhou M, Xu D and Thelen JJ. System analysis of an arabidopsis mutant altered in de novo fatty acid synthesis reveals diverse changes in seed composition and metabolism. Plant Physiology 2009;150(1):27-41.
Brechenmacher L, Lee J, Sachdev S, Song Z, Nguyen THN, Joshi T, Oehrle N, Libault M, Mooney B, Xu D, Cooper B and Stacey G. Establishment of a protein reference map for soybean root hair cells. Plant Physiology 2009;149(2):670-682.
McCaw DL, Chan AS, Stegner AL, Mooney B, Bryan JN, Turnquist SE, Henry CJ, Alexander H and Alexander S. Proteomics of canine lymphoma identifies potential cancer-specific protein markers. Clinical Cancer Research 2007;13(8):2496-2503.
Katz ML, Sanders DN, Mooney BP and Johnson GS. Accumulation of glial fibrillary acidic protein and histone H4 in brain storage bodies of Tibetan terriers with hereditary neuronal ceroid lipofuscinosis. Journal of Inherited Metabolic Disease 2007;30(6):952-963.
Mooney BP, Miernyk JA, Michael Greenlief C and Thelen JJ. Using quantitative proteomics of Arabidopsis roots and leaves to predict metabolic activity. Physiologia Plantarum 2006;128(2):237-250.
Wan J, Torres M, Ganapathy A, Thelen J, DaGue BB, Mooney B, Xu D and Stacey G. Proteomic analysis of soybean root hairs after infection by Bradyrhizobium japonicum. Molecular Plant-Microbe Interactions 2005;18(5):458-467.
Mooney BP and Thelen JJ. High-throughput peptide mass fingerprinting of soybean seed proteins: Automated workflow and utility of UniGene expressed sequence tag databases for protein identification. Phytochemistry 2004;65(12):1733-1744.
Szurmak B, Strokovskaya L, Mooney BP, Randall DD and Miernyk JA. Expression and assembly of Arabidopsis thaliana pyruvate dehydrogenase in insect cell cytoplasm. Protein Expression and Purification 2003;28(2):357-361.
Luethy MH, Gemel J, Johnston ML, Mooney BP, Miernyk JA and Randall DD. Developmental expression of the mitochondrial pyruvate dehydrogenase complex in pea (Pisum sativum) seedlings. Physiologia Plantarum 2001;112(4):559-566.
Mooney BP, Henzl MT, Miernyk JA and Randall DD. The dihydrolipoyl acyltransferase (BCE2) subunit of the plant branched- chain alpha-ketoacid dehydrogenase complex forms a 24-mer core with octagonal symmetry. Protein Science 2000;9(7):1334-1339.
Mooney BP, David NR, Thelen JJ, Miernyk JA and Randall DD. Histidine modifying agents abolish pyruvate dehydrogenase kinase activity. Biochemical and Biophysical Research Communications 2000;267(2):500-503.
- The Plastics, My Friend, Are Blowing in the Wind (St. Louis Riverfront Times, Mar 2009)
- "Green" Plastics Could Help Reduce Carbon Footprint (Feb 2009)
- Researcher 'Grows' Environmentally Friendly Plastic (Feb 2009)
- Utility Patent issued November 7 2000 - Randall D.D., Miernyk J.A., Mooney B.P., Johnston M.L. and Luethy M.H. U.S. Patent No. 6,143,561. DNA encoding plastid pyruvate dehydrogenase and branched chain oxoacid dehydrogenase components.
- University of Missouri System Presidential recognition of academic and public service to the University for Patent award - named one of the “Patent heroes” of the UM system
- Utility Patent issued August 10 2004 - Randall D.D., Mooney B.P., Johnston M.L., Luethy M.H., and Miernyk J.A. U.S. Patent No. 6,773,917. Use of DNA encoding plastid pyruvate dehydrogenase and branched chain oxoacid dehydrogenase components to enhance polyhydroxyalkanoate biosynthesis in plants.