Rubisco proton production can drive the elevation of CO₂ within condensates and carboxysomes.

Abstract

Membraneless organelles containing the enzyme ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) are a common feature of organisms utilizing CO₂ concentrating mechanisms to enhance photosynthetic carbon acquisition. In cyanobacteria and proteobacteria, the Rubisco condensate is encapsulated in a proteinaceous shell, collectively termed a carboxysome, while some algae and hornworts have evolved Rubisco condensates known as pyrenoids. In both cases, CO₂ fixation is enhanced compared with the free enzyme. Previous mathematical models have attributed the improved function of carboxysomes to the generation of elevated CO₂ within the organelle via a colocalized carbonic anhydrase (CA) and inwardly diffusing HCO₃^-, which have accumulated in the cytoplasm via dedicated transporters. Here, we present a concept in which we consider the net of two protons produced in every Rubisco carboxylase reaction. We evaluate this in a reaction-diffusion compartment model to investigate functional advantages these protons may provide Rubisco condensates and carboxysomes, prior to the evolution of HCO₃^- accumulation. Our model highlights that diffusional resistance to reaction species within a condensate allows Rubisco-derived protons to drive the conversion of HCO₃^- to CO₂ via colocalized CA, enhancing both condensate [CO₂] and Rubisco rate. Protonation of Rubisco substrate (RuBP) and product (phosphoglycerate) plays an important role in modulating internal pH and CO₂ generation. Application of the model to putative evolutionary ancestors, prior to contemporary cellular HCO₃^- accumulation, revealed photosynthetic enhancements along a logical sequence of advancements, via Rubisco condensation, to fully formed carboxysomes. Our model suggests that evolution of Rubisco condensation could be favored under low CO₂ and low light environments.