Directions for Optimization of Photosynthetic Carbon Fixation: RuBisCO's Efficiency May Not Be So Constrained After All

Abstract

The ubiquitous enzyme Ribulose 1,5-bisphosphate carboxylase-oxygenase (RuBisCO) fixes atmospheric carbon dioxide within the Calvin-Benson cycle that is utilized by most photosynthetic organisms. Despite this central role, RuBisCO's efficiency surprisingly struggles, with both a very slow turnover rate to products and also impaired substrate specificity, features that have long been an enigma as it would be assumed that its efficiency was under strong evolutionary pressure. RuBisCO's substrate specificity is compromised as it catalyzes a side-fixation reaction with atmospheric oxygen; empirical kinetic results show a trend to tradeoff between relative specificity and low catalytic turnover rate. Although the dominant hypothesis has been that the active-site chemistry constrains the enzyme's evolution, a more recent study on RuBisCO stability and adaptability has implicated competing selection pressures. Elucidating these constraints is crucial for directing future research on improving photosynthesis, as the current literature casts doubt on the potential effectiveness of site-directed mutagenesis to improve RuBisCO's efficiency. Here we use regression analysis to quantify the relationships between kinetic parameters obtained from empirical data sets spanning a wide evolutionary range of RuBisCOs. Most significantly we found that the rate constant for dissociation of CO2 from the enzyme complex was much higher than previous estimates and comparable with the corresponding catalytic rate constant. Observed trends between relative specificity and turnover rate can be expressed as the product of negative and positive correlation factors. This provides an explanation in simple kinetic terms of both the natural variation of relative specificity as well as that obtained by reported site-directed mutagenesis results. We demonstrate that the kinetic behaviour shows a lesser rather than more constrained RuBisCO, consistent with growing empirical evidence of higher variability in relative specificity. In summary our analysis supports an explanation for the origin of the tradeoff between specificity and turnover as due to competition between protein stability and activity, rather than constraints between rate constants imposed by the underlying chemistry. Our analysis suggests that simultaneous improvement in both specificity and turnover rate of RuBisCO is possible.

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

“…This is not very feasible in the context of a likely ancient origin of CCMs in Cyanobacteria. Hence, it raises the possibility that the inherent biochemical and structural constraints might differ not only among different types of RuBisCO, but even within the same type (as observed in form IB from Cyanobacteria versus plants), in response to different evolutionary pressures, leading to a higher RuBisCO kinetic plasticity than previously thought (Cummins et al, 2018). The notable increase in RuBisCO kinetics description of a wide range of species in the last few years allowed us to have a broader picture (see Figure 2).…”

“…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).…”

Revisiting tradeoffs in Rubisco kinetic parameters

“…5,6,22,33,34 If CO 2 bound a specific site on Rubisco before reacting, it might be possible to modulate K C by mutation without substantially affecting the kinetics of subsequent reaction steps. In the unlikely case that gas addition is substantially reversible, 34,35 we might expect to find Rubiscos that evolved enhanced selectivity by energy-coupled kinetic proofreading. Energy coupling can enable amplification of selectivity due to differential CO 2 and O 2 off-rates.…”

“…It is also important to revisit the classic experiments undergirding our understanding of the Rubisco catalytic mechanism, especially those supporting the central assumptions that (i) there is no Michaelis complex for CO 2 or O 2 and (ii) that gas addition is irreversible. 22,34,35 These assumptions substantially constrain CO 2 specificity. If we were to find Rubiscos for which these assumptions are relaxed, they might serve as a basis for engineering a fast-and-selective carboxylase.…”

Crop Biotechnology for Improving Drought Tolerance: Targets, Approaches, and Outcomes

“…Overexpression of Rubisco alone has not given the desired results in C3 rice, already saturated with the enzyme (Suzuki et al, 2007), but recently its overexpression in C4 maize (which naturally contains 70% less rubisco than rice) in concert with Rubisco assembly factor RAF1, has enhanced carbon fixation and productivity (Salesse-Smith et al, 2018) and may pave the way for improving photosynthesis under drought. The development of recombinant Rubisco, without or with decreased oxygenase activity and with higher catalytic rates is so far without success (Bathellier et al, 2018), but increasing the capacity for regeneration of ribulose bisphosphate via overexpression of sedoheptulose-1:7-bisphosphatase in crops (Köhler et al, 2017) remains a promising lead for crop improvement under stress (for more detailed reviews of other targets see Long et al, 2006;Sonnewald and Fernie, 2018;Cummins et al, 2018). Photosynthetic resilience to short-term drought stress has also been evidenced in rice seedlings overexpressing C4 phosphoenolpyruvate carboxylases (PEPCs) from maize and millet (Ding et al, 2015).…”