Lys, pro and trp are critical core amino acid residues recognized by FUM20, a monoclonal antibody against serine protease pan-fungal allergens.
BACKGROUND
Alkaline/vacuolar serine proteases comprise a major group of pan-fungal allergens from several prevalent airborne fungal species. It is of importance to characterize antigenic determinant(s) recognized by monoclonal antibodies against these major allergens.


METHODS
The antigenic determinant of fungal serine proteases recognized by a monoclonal antibody, FUM20, was analyzed by dot immunoassay of synthetic peptides immobilized on cellulose membrane. Results obtained were confirmed by wild-type recombinant protease and its mutants. The epitopes were mapped to the structure of serine proteases by molecular modeling.


RESULTS
A linear epitope encompassing 9 amino acids from Pen ch 18 ((6)EKNAPWGLA(14)) binds FUM20. The corresponding peptide ((5)AKGAPWGLA(13)) from Rho m 2 also binds FUM20. Substitution of K6, P9 or W10 with alanine in this peptide resulted in drastic loss of FUM20 binding. Rho m 2 mutants with single K6A, P9A, P9G, W10A or W10F substitute showed negative immunoblot reactivity against FUM20. However, the Rho m 2 K6R mutant can bind FUM20. Three-dimensional structural models of the FUM20 antigenic determinants on serine proteases were constructed. The lysine residue critical for FUM20 interaction is on the surface of the proteases and solvent accessible. The critical core residue proline is located at the beginning of an alpha-helix.


CONCLUSIONS
The lysine, proline and tryptophan residues located on the N-terminal region of fungal serine proteases are critical core amino acid residues recognized by FUM20, a monoclonal antibody against serine protease pan-fungal allergens. These findings advance our understanding of the antigenic structures responsible for the antigenicity of serine protease allergens.
Lee
Lin-Hau
LH
Department of Medical Research and Education, Taipei Veterans General Hospital, Taipei, Taiwan, ROC.
Tam
Ming F
MF
Chou
Hong
H
Tai
Hsiao-Yun
HY
Shen
Horng-Der
HD
D000485:Allergens / Q000276:immunology; D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000946:Antigens, Fungal / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D001483:Base Sequence; D018604:Epitope Mapping; D005656:Fungal Proteins / Q000276:immunology; D015151:Immunoblotting; D008239:Lysine / Q000276:immunology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011392:Proline / Q000276:immunology*; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000276:immunology; D017386:Sequence Homology, Amino Acid; D012697:Serine Endopeptidases / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D014364:Tryptophan / Q000276:immunology*
D000485:Allergens; D000911:Antibodies, Monoclonal; D000946:Antigens, Fungal; D005656:Fungal Proteins; D011994:Recombinant Proteins; D014364:Tryptophan; D011392:Proline; D012697:Serine Endopeptidases; D008239:Lysine