Binding mechanism of pheromone-binding proteins from the gypsy moth, Lymantria dispar

Pheromone perception in insects plays important roles for their survival and development. They communicate with the environment through sensing and releasing signal molecules, such as host odors or sex pheromones. Pheromone-binding proteins (PBPs) are among the indispensable components of the olfactory system in insects. For example, fly mutants lacking a PBP do not respond to a stimulus of pheromone. PBPs bind pheromone. There are accumulating data on the binding strength and structure. Yet, not until recently has the PBP binding been bridged to its function: a PBP mutant locked in the active conformation found in the PBP.pheromone complex has been shown to stimulate the sensory neurons, even in the absence of pheromone. In this work, I have kinetically dissected the interaction between PBP and ligand, for two PBPs from the gypsy moth, Lymantria dispar: PBP1 and PBP2. Starting with PBP2 and its cognate ligand, (+)-disparlure, I have identified a slow process following a fast interaction. A subsequent study with a fluorescent dye has further identified a diffusion-controlled step prior to a unimolecular relaxation process. Both studies suggest that insect PBPs bind ligand progressively. I have proposed a model that PBP and ligand collide to form an encountered intermediate, P.Lenc, followed by a decay of this species to two stable P.L complexes: external P.Lext and internal P.Lint. Each of these two binding modes of ligand to PBP may correspond to a different function of the PBP. In addition, tryptophan quenching experiments have shown different local conformational changes upon ligand binding. Parallel experiments with truncated PBPs lacking the C-terminal peptide have demonstrated the importance of that segment in PBP function. Efforts have been put to correlate the binding affinity, blend effect and conformation of the LdisPBPs in the presence of synthetic compounds with the effects these compounds have on the electroantennogram (EAG) traces of pheromone stimuli. No correlation was found between affinity of the compounds and short-term/long-term effects of the compounds on the EAG trace. Two subsets of compounds showed a correlation between blend effects and EAG peak broadening. The interpretation is that certain compounds might stabilize the active PBP.L conformation.