ECOFix · Engineering Catalytic Membraneless Organelles for CO2 Fixation

Engineering Catalytic Membraneless Organelles for CO2 Fixation

The accelerating climate crisis urges humanity to understand the global carbon cycle and the biosphere’s capacity to capture and fix atmospheric carbon dioxide. Despite a dramatic rise in CO2 levels from 280 to 420 ppm since the industrial revolution, the effective CO2 concentration in the atmosphere remains low. This presents a fundamental challenge for carbon fixation and carbon capture. To grow at these low concentrations, aquatic phototrophs have evolved so-called carbon-concentrating mechanisms (CCMs) that locally increase CO2 levels in close proximity to Rubisco (Ribulose-1,5-bisphosphate carboxylase oxygenase) for enhancing carbon fixation. Algae, which account for an estimated 30–50% of global carbon fixation, have independently developed two distinct CCMs: cyanobacteria utilize carboxysomes - highly ordered protein microcompartments - while green algae employ membraneless organelles known as pyrenoids. Because Rubisco’s catalytic efficiency is inherently limited by a Pareto optimum between its activity and selectivity for CO2 over O2, with the latter leading to energetically costly photorespiration, CCMs such as the pyrenoid help overcome this limitation by creating microenvironments with elevated CO2, suppressing the wasteful side reaction with O2. Pyrenoids are particularly intriguing because it is crucially formed by just one intrinsically disordered protein (IDP) called EPYC1. In this project we aimed at engineering bottom-up artificial pyrenoids fusion proteins of Rubisco and IDPs that drive liquid-liquid phase separation. We set out with three main objectives: (1) generating artificial pyrenoid-like condensates in vitro and characterizing their structural and catalytic properties, (2) transplanting these artificial organelles into cyanobacteria to replace native carboxysomes and studying their physiological impact, and (3) applying adaptive laboratory evolution under CO2-limiting conditions to optimize these synthetic CCMs for enhanced carbon fixation. This project aims not only to unravel the principles governing pyrenoid assembly and function but also to explore new strategies for improving photosynthetic efficiency in diverse organisms with implications for climate change mitigation and synthetic biology.

Data: CORDIS, © European Union