Three pheromone-binding proteins help segregation between two Helicoverpa species utilizing the same pheromone components

The two sibling species Helicoverpa armigera and Helicoverpa assulta utilise the same two aldehydes as their sex pheromones, but in opposite ratios. In both species three odorant-binding proteins (OBPs) can be classified as pheromone-binding proteins (PBPs). To investigate the role of these three PBPs in chemical communication between sexes and their mode of action, we have expressed the proteins in bacteria and prepared mutants lacking their C-terminal regions. Using polyclonal antibodies we found that the expression of the three PBPs is basically confined to the antennae of both sexes and both species. Binding experiments with the fluorescent probe N-phenyl-1-naphthylamine across a pH range indicated that, the affinity of wild-type proteins decreases at low pH, while that of the mutants is not or less affected, suggesting that a conformational change of the C-terminus occurs in these proteins, as reported for other lepidopteran OBPs. All three proteins bind with similar strength both pheromone components, as well as their corresponding alcohols and acetates. However, they exhibit significant selectivity to linear alcohols and aldehydes of different length, with optimal affinities to the ligand of 13–15 carbon atoms for PBP1 and 12–14 carbon atoms for PBP2. We suggest that all three PBPs might cooperate to build a unique olfactory image, that could help avoiding cross-mating between the two species and with other noctuids.

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► Two sibling species Helicoverpa armigera and Helicoverpa assulta express three PBPs specifically in antennae of both sexes.
► The three PBPs show poor discrimination between functional groups, but good selectivity to molecular size of ligands.
► Binding is affected by pH in different fashions for the three PBPs and their C-terminal truncated mutants.
► Three PBPs might contribute to build a unique olfactory image of the pheromonal blend, thus avoiding interspecific mating.