Investigator Lead: Prof. Mike Blatt
Participating Laboratories: Glasgow, Penn State (Golbeck and Kumar), Warwick, Berkeley, MSU
We are pursuing two strategies for light-driven HCO3– transport. The first is to introduce pHR and a HCO3–/Cl- antiporter into the membrane. The idea is to use the light-driven Cl- pump of halorhodopsin (pHR) to accumulate Cl- and generate a secondary driving force for exchange of Cl- with HCO3–. The second is to engineer pHR to pump HCO3– directly.
Halorhodopsin is an integral membrane protein found in the salt-tolerant archaeon Halobacterium salinarum. Photoexcitation of the pHR energizes Cl– transport. Ion selectivity is a localized feature of the pHR transport site, making the engineering of a light-driven HCO3– pump a real possibility.
We have established pHR activity following heterologous expression and are developing stopped flow systems to monitor directly its coupling with Cl-/HCO3– exchange. Both proteins can be expressed also in the chloroplast envelope membrane. We have used multi-cistronic vectors for co-expression and targeting, and have localised expression with fluorescent tags and confocal fluorescence microscopy.
Modeling light-driven ion fluxes and on CO2 delivery has uncovered several design challenges and opportunities. Bicarbonate pumps operating at 10 Hz at a density of 60000 µm-2 on the chloroplast membrane are predicted to increase assimilation by 5-18%.