Abstract
Many photosynthetic organisms employ a CO2 concentrating mechanism (CCM) to increase the rate of CO2 fixation via the Calvin cycle. CCMs catalyze ≈50% of global photosynthesis, yet it remains unclear which genes and proteins are required to produce this complex adaptation. We describe the construction of a functional CCM in a non-native host, achieved by expressing genes from an autotrophic bacterium in an Escherichia coli strain engineered to depend on rubisco carboxylation for growth. Expression of 20 CCM genes enabled E. coli to grow by fixing CO2 from ambient air into biomass, with growth in ambient air depending on the components of the CCM. Bacterial CCMs are therefore genetically compact and readily transplanted, rationalizing their presence in diverse bacteria. Reconstitution enabled genetic experiments refining our understanding of the CCM, thereby laying the groundwork for deeper study and engineering of the cell biology supporting CO2 assimilation in diverse organisms.
Highlights
Most carbon in the biosphere enters by CO2 fixation in the Calvin-Benson-Bassham cycle (Bassham, 2003; Bassham et al, 1954; Benson, 2002; Field et al, 1998; Raven, 2009)
Using a genome-wide screen in the CO2-fixing proteobacterium Halothiobacillus neapolitanus, we recently demonstrated that a 20-gene cluster encodes all activities required for the concentrating mechanism (CCM), at least in principle (Desmarais et al, 2019)
In order to evaluate the effect of heterologous CCM expression, we first designed an E. coli strain that depends on rubisco carboxylation for growth
Summary
Most carbon in the biosphere enters by CO2 fixation in the Calvin-Benson-Bassham cycle (Bassham, 2003; Bassham et al, 1954; Benson, 2002; Field et al, 1998; Raven, 2009). Ribulose Bisphosphate Carboxylase/Oxygenase - commonly known as rubisco - is the CO2 fixing enzyme in this cycle (Kawashima and Wildman, 1971; Weissbach et al, 1956; Wildman, 2002) and likely the most abundant enzyme on Earth (Bar-On and Milo, 2019). All known rubiscos catalyze a competing oxygenation of the five-carbon organic substrate, ribulose 1, 5-bisphosphate (Bathellier et al, 2018; Bowes and Ogren, 1972; Cleland et al, 1998). Rubisco oxygenation represents a ‘waste’ of cellular resources on two fronts: it fails to generate any new organic carbon and produces a molecule (2-phosphoglycolate) that is not part of the Calvin cycle and must be recycled through a salvage pathway to keep the cycle going (Busch, 2020)
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