Kinetics of Interactions between LOV Domains from Chlamydomonas Reinhardtii

Kinetics of Interactions between LOV domains from Chlamydomonas reinhardtiiCarey K. Johnson, Kathrin Magerl, Katee Wyant, Ashley McDade, Will Newhart, David C. Arnett, and Bernhard Dick.We have investigated the kinetics of interactions between Light-Oxygen-Voltage (LOV) domains of the light-sensing protein phototropin from Chlamydomonas reinhardtii. The photochemical response of phototropin to blue light involves adduct formation between a flavin mononucleotide cofactor and a nearby Cys residue. Interactions between LOV domains, either domains LOV1 and LOV2 within the same phototropin, or between LOV domains from different phototropin molecules, are thought to play a role in the subsequent activation of the phototropin kinase domain. We studied the kinetics of exchange of LOV domains between complexes (dimers or higher order oligomers) by stop-flow FRET measurements. A biphasic response with time constants from tens to hundreds of seconds was sensitive to prior exposure of LOV domains to blue light. LOV-LOV interactions have also been detected at the single-molecule level by TIRF imaging, providing further information about the interaction kinetics.