Investigation of Thermodynamic Dissociation Kinetics to Determine the Binding Strengths within a Membrane Protein Complex: Photosystem II Supercomplex

Abstract

Protein-protein interactions as well as the formations of protein complexes have been intensively studied by various methods in both in vivo and in vitro. However, although membrane proteins have important roles in cells, interactions within membrane protein complexes have been hardly studied than that within soluble proteins due to the lack of suitable methods, though one approach using mass spectrometry has been recently developed to elucidate membrane protein complexes. Here, we present an approach to determine binding strengths within a membrane protein complex by investigating the thermodynamic dissociation kinetics of an isolated membrane protein complex using time-resolved fluorescence spectroscopy. We demonstrated different bindings of light-harvesting complex II (LHCII) trimers in a photosystem II (PSII) supercomplex isolated from a green alga Chlamydomonas reinhardtii, where the different binding types of the LHCII trimers have been indirectly predicted by the presence or absence of LHCII trimers in negatively stained PSII supercomplex particles observed by electron microscopy but have not been determined. As a result of analysis on thermodynamic dissociation kinetics, we identified three different activation energies for the dissociations of LHCII trimers, which imply the presence of multiple hydrogen bonds and/or ion-dipole bonds between LHCII trimers and core complexes.