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Team:SCUT/Project/System Construction/Co2 Fixation
From 2014.igem.org
Introduction
Having utilizing the high-concentration ATP and CO2 around mitochodria as the starting point, we did research on both endogenous metabolic pathways of S.cerevisiae and exogenous pathways that consuming CO2 and, finally set employing PRK ( phosphoribulokinase ), RuBisCo ( Ribulose-1,5-bisphosphate carboxylase ) and CA ( carbonic anhydrase ) to fix CO2 to improve the C sequestration of yeast，in other words, to increase the yield of pyruvate for butanol production as the standing point. To make full use of ATP and CO2, we tried to find out a leading peptide for locating and exogenous scaffold proteins to fix the above-mentioned enzymes to a limited space. Finally we employed Tom22, a leading peptide of outer mitochondrial membrane, and GBD, SH3 and PDZ and their ligands, to reach that goal.
Design
Construction
Leading Peptide Test
References
[1] Hillel K. Brandes‡, Fred C. Hartman§¶, Tse-Yuan S. Lu§, et al. : Efficient Expression of the Gene for Spinach Phosphoribulokinase in Pichia pastoris and Utilization of the Recombinant Enzyme to Explore the Role of Regulatory Cysteinyl Residues by Site-directed Mutagenesis*. The Journal of Biological Chemistry 1996 Vol. 271, No. 11, Issue of March 15, pp. 6490–6496.
[2] F. Robert Tabita, Sriram Satagopan, Thomas E. Hanson, et al. : Distinct form I, II, III, and IV Rubisco proteins from the three kingdoms of life provide clues about Rubisco evolution and structure/function relationships. Journal of Experimental Botany 2008 Vol. 59, No. 7, pp. 1515–1524.
[3] Christopher R. Somerville and Shauna C. Somerville: Cloning and Expression of the Rhodospirillum rubrum Ribulosebisphosphate Carboxylase Gene in E. coil. Mol Gen Genet 1984 193:214-219
[4] Víctor Guadalupe-Medina, H Wouter Wisselink, Marijke AH Luttik, et al. : Carbon dioxide fixation by Calvin-Cycle enzymes improves ethanol yield in yeast. Biotechnology for Biofuels 2013 6:125.
[5] Walter Bonaccia, Poh K. Tengb, Bruno Afonsoa, et al. : Modularity of a carbon-fixing protein organelle. PNAS 2012 vol. 109 , no. 2: 478-483.
[6] Jian Qiu, Lena-Sophie Wenz, Ralf M. Zerbes, et al. : Coupling of Mitochondrial Import and Export Translocases by Receptor-Mediated Supercomplex Formation. Cell 2013 154, 596–608.
[7] Birgitta M. GEIER', IIermann SCHAGGER', Claus ORTWEIN", et al. : Kinetic properties and ligand binding of the eleven-subunit cytochrome-c oxidase from Saccharomyces cerevisiae isolated. Eur. J. Biochem. 227, 296-302 (1995). Eur. J. Biochem 1995 227, 296-302.
[8] MA Jun, SUN Fei: Translocation of Mitochondrial Proteins. ACTA BIOPHYSICA SINICA 2010 Vol.26 No.10: 880-893
[9] John E Dueber, Gabriel C Wu, G Reza Malmirchegini, et al. : Synthetic protein scaffolds provide modular control. NATURE BIOTECHNOLOGY 2009 Vol. 27 No. 8
[10] Matthew C. Good, Jesse G. Zalatan, Wendell A. Lim† : Scaffold Proteins: Hubs for Controlling the Flow of Cellular. Science 2011 332, 680.
[11] Tae SeokMoona, JohnE.Dueber, EricShiue, et al. : Use of modular, synthetic scaffolds forim proved production of glucaricacid in engineered E. coli over metabolic flux. Metabolic Engineering 2010 12 298–305.
