Extrusion of the C-terminal helix in navel orangeworm moth pheromone-binding protein (AtraPBP1) controls pheromone binding.
The navel orangeworm, Amyelois transitella (Walker), is an agricultural insect pest that can be controlled by disrupting male-female communication with sex pheromones, a technique known as mating disruption. Insect pheromone-binding proteins (PBPs) provide fast transport of hydrophobic pheromones through aqueous sensillar lymph and promote sensitive delivery of pheromones to receptors. Here we present a mutational analysis on a PBP from A. transitella (AtraPBP1) to evaluate how the C-terminal helix in this protein controls pheromone binding as a function of pH. Pheromone binds tightly to AtraPBP1 at neutral pH, but the binding is much weaker at pH below 5. Deletion of the entire C-terminal helix (residues 129-142) causes more than 100-fold increase in pheromone-binding affinity at pH 5 and only a 1.5-fold increase at pH 7. A similar pH-dependent increase in pheromone binding is also seen for the H80A/H95A double mutant that promotes extrusion of the C-terminal helix by disabling salt bridges at each end of the helix. The single mutants (H80A and H95A) also exhibit pheromone binding at pH below 5, but with ∼2-fold weaker affinity. NMR and circular dichroism data demonstrate a large overall structural change in each of these mutants at pH 4.5, indicating an extrusion of the C-terminal helix that profoundly affects the overall structure of the low pH form. Our results confirm that sequestration of the C-terminal helix at low pH as seen in the recent NMR structure may serve to block pheromone binding. We propose that extrusion of these C-terminal residues at neutral pH (or by the mutations in this study) exposes a hydrophobic cleft that promotes high affinity pheromone binding.
Department of Entomology, University of California, Davis, CA 95616, USA.
Wei
W
Xu
Xianzhong
X
Xu
Walter S
WS
Leal
James B
JB
Ames
D000818:Animals; D002352:Carrier Proteins / Q000737:chemistry* / Q000235:genetics; D002942:Circular Dichroism; D004252:DNA Mutational Analysis; D005260:Female; D006863:Hydrogen-Ion Concentration; D019476:Insect Proteins / Q000737:chemistry* / Q000235:genetics; D008297:Male; D009036:Moths / Q000378:metabolism*; D009154:Mutation; D019906:Nuclear Magnetic Resonance, Biomolecular; D010675:Pheromones / Q000737:chemistry*; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary
D002352:Carrier Proteins; D019476:Insect Proteins; D010675:Pheromones; C403713:pheromone binding protein, insect