Characterization of a serine protease homologous to house dust mite group 3 allergens from the scabies mite Sarcoptes scabiei.
The scabies mite, Sarcoptes scabiei var. hominis, infests human skin, causing allergic reactions and facilitating bacterial infection by Streptococcus sp., with serious consequences such as rheumatic fever and rheumatic heart disease. To identify a possible drug target or vaccine candidate protein, we searched for homologues of the group 3 allergen of house dust mites, which we subsequently identified in a cDNA library. The native protein, designated Sar s 3, was shown to be present in the mite gut and excreted in fecal pellets into mite burrows within the upper epidermis. The substrate specificity of proteolytically active recombinant rSar s 3 was elucidated by screening a bacteriophage library. A preference for substrates containing a RS(G/A) sequence at the P1-P2' positions was revealed. A series of peptides synthesized as internally quenched fluorescent substrates validated the phage display data and high performance liquid chromatography/mass spectrometry analysis of the preferred cleaved substrate and confirmed the predicted cleavage site. Searches of the human proteome using sequence data from the phage display allowed the in silico prediction of putative physiological substrates. Among these were numerous epidermal proteins, with filaggrin being a particularly likely candidate substrate. We showed that recombinant rSar s 3 cleaves human filaggrin in vitro and obtained immunohistological evidence that the filaggrin protein is ingested by the mite. This is the first report elucidating the substrate specificity of Sar s 3 and its potential role in scabies mite biology.
Beckham
Simone A
SA
Infectious Diseases and Immunology Division, Queensland Institute for Medical Research, Herston, Queensland 4029, Australia.
Boyd
Sarah E
SE
Reynolds
Simone
S
Willis
Charlene
C
Johnstone
Masego
M
Mika
Angela
A
Simerská
Pavla
P
Wijeyewickrema
Lakshmi C
LC
Smith
A Ian
AI
Kemp
David J
DJ
Pike
Robert N
RN
Fischer
Katja
K
D000818:Animals; D039741:Antigens, Dermatophagoides / Q000737:chemistry*; D001435:Bacteriophages / Q000378:metabolism; D005260:Female; D000091344:Filaggrin Proteins; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007381:Intermediate Filament Proteins / Q000737:chemistry; D051379:Mice; D008810:Mice, Inbred C57BL; D008925:Mites; D019151:Peptide Library; D010843:Pichia / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry; D012520:Sarcoptes scabiei / Q000737:chemistry*; D057057:Serine Proteases / Q000737:chemistry*; D013379:Substrate Specificity
D039741:Antigens, Dermatophagoides; C000717249:FLG protein, human; D000091344:Filaggrin Proteins; D007381:Intermediate Filament Proteins; D019151:Peptide Library; D011994:Recombinant Proteins; D057057:Serine Proteases