Evolving Methanococcoides burtonii archaeal Rubisco for improved photosynthesis and plant growth

Abstract

In photosynthesis Ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) catalyses the often rate limiting CO2-fixation step in the Calvin cycle. This makes Rubisco both the gatekeeper for carbon entry into the biosphere and a target for functional improvement to enhance photosynthesis and plant growth. Encumbering the catalytic performance of Rubisco is its highly conserved, complex catalytic chemistry. Accordingly, traditional efforts to enhance Rubisco catalysis using protracted “trial and error” protein engineering approaches have met with limited success. Here we demonstrate the versatility of high throughput directed (laboratory) protein evolution for improving the carboxylation properties of a non-photosynthetic Rubisco from the archaea Methanococcoides burtonii. Using chloroplast transformation in the model plant Nicotiana tabacum (tobacco) we confirm the improved forms of M. burtonii Rubisco increased photosynthesis and growth relative to tobacco controls producing wild-type M. burtonii Rubisco. Our findings indicate continued directed evolution of archaeal Rubisco offers new potential for enhancing leaf photosynthesis and plant growth.

Evolutionary trends in RuBisCO kinetics and their co‐evolution with CO2 concentrating mechanisms

“…The analyzed archaeal species are anaerobic organisms never facing O 2 in their environment, and whose RuBisCO is not involved in autotrophic carbon fixation but in the assimilation of ribonucleosides. Most notably, half‐saturation constants for CO 2 ( K c ) and O 2 ( K o ) for RuBisCOs from the three archaeal species analyzed to date (Kreel and Tabita, , ; Wilson et al , ) share a common trend not previously observed in any other RuBisCOs. They all possess a stronger RuBisCO affinity for O 2 than for CO 2 at their optimum growth temperature (83°C for the hyperthermophilic species Thermococcus kodakarensis and Archaeoglobus fulgidus and 25°C for the Antarctic species M. burtonii ), which might mirror the kinetics of the common ancestor bona fide RuBisCO with an anoxygenic and probably heterotrophic origin.…”

“…During this time, substantial progress has been made in understanding the structure of the active site and the complete reaction mechanism catalyzed by RuBisCO (Andersson, 2008). Although most of the studies on RuBisCO structural properties and its reaction mechanism have been focused on a few model species, and large-scale explorations of RuBisCO catalytic traits have been frequently restricted to angiosperm species (Galm es et al, 2005;Kubien et al, 2008;Hermida-Carrera et al, 2016;Orr et al, 2016;Sharwood et al, 2016a), recent research studies have shed light on the variability of biochemical and molecular RuBisCO traits in previously underreported groups (Satagopan et al, 2014;Galm es et al, 2014aGalm es et al, , 2015Galm es et al, , 2016Wilson et al, 2016;Young et al, 2016;Heureux et al, 2017;Valeg ard et al, 2018;Iñiguez et al, 2018). The increase in the availability of RuBisCO measurements on phylogenetically distant groups have enabled a more profound analysis of RuBisCO fine tuning through evolution (Liu et al, 2017;Young and Hopkinson, 2017;Cummins et al, 2018;Tcherkez et al, 2018).…”

RubisCO selection using the vigorously aerobic and metabolically versatile bacterium Ralstonia eutropha

“…There has been considerable interest in using directed evolution strategies to improve the expression levels, assembly and kinetic performance of RubisCO so that the rates for net carboxylation may be maximized [5,[7][8][9][10][11][12][14][15][16][17][36][37][38]. The presence of an efficient lithoautotrophic metabolism has attracted recent interest in tapping R. eutropha for biotechnological CO 2 utilization [3,4,39,40].…”

Surveying Rubisco diversity and temperature response to improve crop photosynthetic efficiency

“…However, advances in engineering precise changes in model systems continue to provide important developments that are increasing our understanding of Rubisco catalysis (Spreitzer et al, 2005;Whitney et al, 2011aWhitney et al, , 2011bMorita et al, 2014;Wilson et al, 2016), regulation (Andralojc et al, 2012;Carmo-Silva and Salvucci, 2013;Bracher et al, 2015), and biogenesis (Saschenbrecker et al, 2007;Whitney and Sharwood, 2008;Lin et al, 2014b;Hauser et al, 2015;Whitney et al, 2015).…”