Faecally Derived Hydrolytic Enzymes from <i>Dermatophagoides pteronyssinus</i>: Physicochemical Characterisation of Potential Allergens

Abstract

The previous findings that the group I and III mite allergens, and amylase present in mite faeces are hydrolytic enzymes has prompted a study to determine whether this material contains other enzymes which could be allergenic. Thus, spent growth medium devoid of whole Dermatophagoides pteronyssinus mites was shown to contain glucoamylase, lipase and lysozyme in addition to the cysteine protease, serine protease and amylase activities associated with the above allergens, respectively. All of these enzymes are probably associated with mite digestive processes. They were rapidly solubilised, heterogeneous with regard to charge (pI in the range 4–8) and demonstrated maximum biochemical activity in the neutral pH range. Three serine proteases were detected and comprised a chymotrypsin-like, a trypsin-like and an unclassified enzyme with pI of 4.1 and 5.3, 8.5 and 7.1, respectively. Only one cysteine protease was observed, which paralleled immunochemically identified Der p I in a variety of assays. It was shown to cleave at lysyl residues and could be inhibited by the specific cysteine protease inhibitor, E-64. The remaining serine proteases, glucoamylase, lipase and lysozyme represent potential allergens.

Allergenicity and Physicochemical Characterization of House Dust Mite Derived Amylase

“…Enzyme activity was easily detected in extracts of whole mite and spent growth medium (which was devoid of whole mites) but not in unused growth medium, suggesting it was present in faecal pellets [4]. Solubility data, which showed that the en zyme eluted from dry medium within 40 s after con tact with an aqueous solvent, were consistent with this proposal [5]. Amylase was also found in 20 out of 20 extracts of dust obtained from mattresses (geometric mean 1.95; range 0.6-15.3 units/g fine dust) and in 18 out of 20 from lounge room carpets (geometric mean 1.83; range 0.6-4.4 units/g fine dust).…”

Der p 1 facilitates transepithelial allergen delivery by disruption of tight junctions

“…Der p 1 was purified as described (27,28) to >98% ( Figure 1). Enzymatic activity was measured spectrophotometrically using N-benzoyl-FVR-p-nitroanilide (single-letter amino acid notation).…”

Crucial Commitment of Proteolytic Activity of a Purified Recombinant Major House Dust Mite Allergen Der p1 to Sensitization toward IgE and IgG Responses

“…Group 1 and group 2 allergens are major allergens derived from house dust mites based on the frequency of patients sensitized, amount of specific IgE, and content in mite extract (4 -10). The group 1 allergens, Der p 1 from D. pteronyssinus and Der f1 from D. farinae, belong to the papain-like cysteine protease family (11,12) and actually exhibit cysteine protease activity (13)(14)(15)(16). The cDNAs for Der p 1 and Der f 1 were isolated when cDNA cloning of allergens began (11,12,17).…”