The Interaction of Rubisco and Rubisco Activase: A FRET-based Study

Rubisco is a very important protein which catalyzes the addition of CO2 to ribulose-1,5-bisphosphate (RuBP) to form two molecules of 3-phosphoglycerate in photosynthesis. Rubisco activase is the protein which functions to uninhibit Rubisco, however proof of a physical interaction has never been shown. A possible method for determining the interaction of the two proteins is by Förster Resonance Energy Transfer (FRET) based analysis of the two proteins. Attempts to get a FRET signal from these two proteins have been unsuccessful. To get better results, Ficoll 70, a crowding agent, was used. Analysis suggests that Ficoll 70 does not affect the fluorescence of Alexa-fluor 488 and Alexa-fluor 647 used to label the two proteins. Further analysis also suggests that while the Alexa label on Rubisco activase does not affect the ATPase activity of the protein, the protein also does not have a high rate of ATP turnover.