Comparative modelling of major house dust mite allergen <i>Der p</i> I: structure validation using an extended environmental amino acid propensity table

Abstract

A model of the 3-D structure of a major house dust mite allergen Der p I associated with hypersensitivity reactions in humans was built from its amino acid sequence and its homology to three known structures, papain, actinidin and papaya proteinase omega of the cysteine proteinase family. Comparative modelling using COMPOSER was used to arrive at an initial model. This was refined using interactive graphics and energy minimization with the AMBER force field incorporated in SYBYL (Tripos Associates). Compatibility of the Der p I amino acid sequence with the cysteine proteinase fold was checked using an environment-dependent amino acid propensity table incorporated into a new program HARMONY with a variable length windowing facility. A five-residue window was used to probe local conformational integrity. Propensities were derived from a structural alignment database of homologous proteins using a robust entropy-driven smoothing procedure. Der p I shares essential structural and mechanistic features with other papain-like cysteine proteinases, including cathepsin B. The active-site thiolate-imidazolium ion pair comprises the side chains of Cys34 and His170. A cystine disulfide not present in other known structures bridges residue 4 of an N-terminal extension and the core residue 117. Two conserved disulfide bridges are formed by residues 31 and 71 and residues 65 and 103. Model building of peptide substrate analogue complexes suggests a preference for phenylalanyl or basic residues at the P2 position, whilst selectivity may be of minor importance at the S1 subsite. The electrostatic influences on the Der p I active-site ion pair and extended peptide binding region are markedly different from those in known structures. A highly immunogenic surface exposed region (residues 107-131), comprising several overlapping T cell epitope sites, has no shared sequence identity with human liver cathepsin B and contains three insertion-deletion sites. The structure provides a basis for testing the substrate specificity of Der p I and the potential role of proteinase activity in hypersensitivity reactions. These studies may offer a new treatment strategy by hyposensitization with inactive mutants or mutants with significantly altered proteinase activity, either alone or complexed with antibody.