Hydrophobic ligands influence the structure, stability, and processing of the major cockroach allergen Bla g 1

The cockroach allergen Bla g 1 forms a novel fold consisting of 12 amphipathic alpha-helices enclosing an exceptionally large hydrophobic cavity which was previously demonstrated to bind a variety of lipids. Since lipid-dependent immunoactivity is observed in numerous allergens, understanding the structural basis of this interaction could yield insights into the molecular determinants of allergenicity. Here, we report atomic modelling of Bla g 1 bound to both fatty-acid and phospholipids ligands, with 8 acyl chains suggested to represent full stoichiometric binding. This unusually high occupancy was verified experimentally, though both modelling and circular dichroism indicate that the general alphahelical structure is maintained regardless of cargo loading. Fatty-acid cargoes significantly enhanced thermostability while inhibiting cleavage by cathepsin S, an endosomal protease essential for antigen processing and presentation; the latter of which was found to correlate to a decreased production of known T-cell epitopes. Both effects were strongly dependent on acyl chain length, with 18-20 carbons providing the maximal increase in melting temperature (similar to 20 degrees C) while completely abolishing proteolysis. Diacyl chain cargoes provided similar enhancements to thermostability, but yielded reduced levels of proteolytic resistance. This study describes how the biophysical properties of Bla g 1 ligand binding and digestion may relate to antigen processing, with potential downstream implications for immunogenicity.