New tools for identifying nature's solutions for improving CO<sub>2</sub> fixation — ASN Events
  1. Schedule
  2. Session Five - Poster Session A
  3. New tools for identifying nature's solutions for improving CO2 fixation

New tools for identifying nature's solutions for improving CO2 fixation (#147)

Laura H Gunn 1 , Rosemary J Birch 1 , Robert E Sharwood 2 , Spencer M Whitney 1
  1. Australian National University, Canberra, ACT, Australia
  2. University of Western Sydney, Sydney, NSW, Australia

The activity of the CO2-fixing enzyme Ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) has a pervasive influence on photosynthetic carbon assimilation, often limiting plant growth.  Despite millions of years of evolution, Rubisco is considered an inefficient enzyme as O2 competes with CO2 as a substrate producing a product (2-phosphoglycolate).  Recycling 2-phosphoglycolate via photorespiration consumes energy and releases fixed CO2.  However, Rubisco exhibits substantial natural catalytic diversity particularly among the most structurally complex L8S8 form I Rubiscos found in plants, algae, cyanobacteria and proteobacteria.  This Rubisco form comprises a core of eight large (L) subunits, harbouring 8 catalytic sites, and tetramers of small (S) subunits capping each end.  The use of foreign Rubiscos with favourable kinetic properties, chimeric Rubisco complexes, or endogenous Rubisco with specific mutations that enhance its catalytic rate could improve photosynthetic carbon assimilation in plants, particularly in key grain crops123.  However, a major limitation to identifying solutions for improving crop Rubisco catalysis is that the folding and assembly requirements of most L8S8 Rubiscos can not be met by high throughput expression systems such as E.coli. Structure-function studies of naturally more efficient Rubiscos thus relies on transplastomic genetic approaches to engineer recombinant Rubisco in leaf chloroplasts.  Current methods for evaluating folding, assembly, and kinetics of recombinant Rubisco in chloroplasts involve biolistic plastome transformation using a custom designed tobacco line called cmtrL14.  Unfortunately the 5 to 12 month timeline required for recombinant Rubisco analysis by this stable transformation approach limits screening throughput.  A new chloroplast transient expression platform technology has been developed to overcome this limitation.  Details of this technology and demonstration of its versatility to examine assembly and catalytic screening of recombinant Rubisco in chloroplasts within a 7 day time frame will be presented.

  1. Long, S.P., Zhu, X.-G., Naidu, S.L. and Ort, D.R. (2006) Can improvement in photosynthesis increase crop yields? Plant Cell Environ. 29, 315-330.
  2. Whitney, S.M., Sharwood, R.E., Orr, D., White, S.J., Alonso, H. and Galmés, J. (2011) Isoleucine 309 acts as a C4 catalytic switch that increases ribulose-1,5-bisphosphate carboxylase/oxygenase (rubisco) carboxylation rate in Flaveria. Proc. Natl. Acad Sci. Unit. States Am. 108, 14688-14693.
  3. Ishikawa, C., Hatanaka, T., Misoo, S., Miyake, C. and Fukayama, H. (2011) Functional Incorporation of Sorghum Small Subunit Increases the Catalytic Turnover Rate of Rubisco in Transgenic Rice. Plant Physiol. 156, 1603-1611.
  4. Whitney, S.M. and Sharwood, R.E. (2008) Construction of a tobacco master line to improve Rubisco engineering in chloroplasts. J. Exp. Bot. 59, 1909-1921.