The mechanism underlying OBP heterodimer formation and the recognition of odors in Holotrichia oblita Faldermann

Insect odorant-binding proteins (OBPs) play key roles in transport odors to receptors and contribute to insect survival. The cooperative interaction of HoblOBP1 and HoblOBP2 in Holotrichia oblita Faldermann (Coleoptera: Melolonthidae) could increase their binding capacity for ligands. In present study, molecular docking results showed that OBP1/OBP2 complex formed a large binding pocket and interacted with the ligands by hydrogen bonds and hydrophobic interactions. Then, nine amino acids for single site mutations, three paired for double sites, and negative control were mutated into alanine successfully by site-directed mutagenesis. Finally, fluorescence binding assays of these mutants showed that breaking one or two pairs of hydrogen bonds between HoblOBP1 and HoblOBP2 or formed with the ligands significantly decrease the binding affinity with the ligands. However, hydrophobic site mutants still showed slight binding affinity to the ligands. Therefore, the three pairs of hydrogen bonds involved in heterodimer formation and the five hydrogen bonding sites in binding pocket played a key role in response to odors in H. oblita. Our findings may promote further understanding of the mechanisms underlying OBP dimer formation and the role of OBP dimers in odor perception and discrimination.