Two Minus-C odorant binding proteins from Helicoverpa armigera display higher ligand binding affinity at acidic pH than neutral pH

Odorant-binding proteins (OBPs), widely and abundantly expressed in insect olfactory organs, are important for insect olfaction. In this study, two novel Minus-C OBP genes (HarmOBP17 and HarmOBP18) were characterized from the cotton bollworm, Helicoverpa armigera, one of the most important pests in cotton and other crops. Quantitative RT-PCR results indicate that the expression levels of the two genes differed greatly in regard to developmental stages, tissues, and genders. Binding properties of the two OBPs with plant volatiles were investigated at different pH levels using a competitive binding assay. Both OBPs displayed the highest binding affinities (Ki < 13 μM) with Beta-ionone among the 85 tested compounds. Surprisingly, the two OBPs (HarmOBP17 in particular) displayed higher binding at pH 5.0 than at pH 7.4 and 10.0. This is the first reported Lepidopteran OBPs showing a contrary pH-dependent ligand binding behavior to other OBPs, implying a different mechanism of ligand binding and release. In addition, binding assays of the mutant OBPs (with the C-terminus eliminated) showed that the C-terminus was important for the ligand binding, but this importance varied with different ligands and OBPs, suggesting that their binding properties depend on the specific interactions between OBP and ligand.