TY - JOUR
T1 - The 18-kDa form of cat allergen Felis domesticus 1 (Fel d 1) is associated with gelatin- and fibronectin-degrading activity.
AU - Ring, P C
AU - Wan, H
AU - Schou, C
AU - Kroll Kristensen, A
AU - Roepstorff, P
AU - Robinson, C
PY - 2000/8/1
Y1 - 2000/8/1
N2 - BACKGROUND: Fel d 1, an important allergen from domestic cats, is a significant cause of asthma. In addition to directly promoting IgE synthesis, other biological activities of allergens may contribute to either allergic sensitization or the magnitude of allergic effector responses. For example, allergens that degrade proteins have been suggested to facilitate allergen presentation by increasing parallelular permeability of airways epithelium. However, little information exists to indicate whether Fel d 1 has other activities relevant to allergic responses. OBJECTIVE: To study whether Fel d 1 is associated with enzyme activity. METHODS: Fel d 1 was obtained by a rigorous purification strategy and its identity confirmed by laser desorption mass spectrometry, cleavage and sequencing. The ability of Fel d 1 to degrade gelatin, fibronectin and the artificial substrate N-benzoyl-FVR-p-nitroanilide was studied. The effect of Fel d 1 on the morphology of tight junctions in epithelial cell monolayers was also investigated. RESULTS: The 18-kDa form of Fel d 1 caused degradation of denatured collagens (gelatin) and cleaved a 20-kDa fragment from the A chain of plasma fibronectin. Catalytic activity was not altered by inhibitors of cysteine peptidases, matrix metallopeptidases or by removal of divalent cations. In contrast, aprotinin and TLCK were inhibitors of Fel d 1. The absence of a serine peptidase catalytic triad in Fel d 1, together with the stoichiometry of the inhibition of TLCK and aprotinin, suggest that their inhibitory action may be due to noncatalytic site interactions. Alternatively, highly purified Fel d 1 may be associated with an active contaminant, although none were found. CONCLUSION: These results suggest that Fel d 1 is another example of a domestic allergen which is associated with enzyme activity. It remains to be established whether the activity resides in Fel d 1 itself or in an unresolved, and possibly related, protein.
AB - BACKGROUND: Fel d 1, an important allergen from domestic cats, is a significant cause of asthma. In addition to directly promoting IgE synthesis, other biological activities of allergens may contribute to either allergic sensitization or the magnitude of allergic effector responses. For example, allergens that degrade proteins have been suggested to facilitate allergen presentation by increasing parallelular permeability of airways epithelium. However, little information exists to indicate whether Fel d 1 has other activities relevant to allergic responses. OBJECTIVE: To study whether Fel d 1 is associated with enzyme activity. METHODS: Fel d 1 was obtained by a rigorous purification strategy and its identity confirmed by laser desorption mass spectrometry, cleavage and sequencing. The ability of Fel d 1 to degrade gelatin, fibronectin and the artificial substrate N-benzoyl-FVR-p-nitroanilide was studied. The effect of Fel d 1 on the morphology of tight junctions in epithelial cell monolayers was also investigated. RESULTS: The 18-kDa form of Fel d 1 caused degradation of denatured collagens (gelatin) and cleaved a 20-kDa fragment from the A chain of plasma fibronectin. Catalytic activity was not altered by inhibitors of cysteine peptidases, matrix metallopeptidases or by removal of divalent cations. In contrast, aprotinin and TLCK were inhibitors of Fel d 1. The absence of a serine peptidase catalytic triad in Fel d 1, together with the stoichiometry of the inhibition of TLCK and aprotinin, suggest that their inhibitory action may be due to noncatalytic site interactions. Alternatively, highly purified Fel d 1 may be associated with an active contaminant, although none were found. CONCLUSION: These results suggest that Fel d 1 is another example of a domestic allergen which is associated with enzyme activity. It remains to be established whether the activity resides in Fel d 1 itself or in an unresolved, and possibly related, protein.
KW - Allergens
KW - Animals
KW - Cats
KW - Chromatography, Affinity
KW - Chromatography, High Pressure Liquid
KW - Collagen
KW - Epithelium
KW - Fibronectins
KW - Gelatin
KW - Glycoproteins
KW - Immunoelectrophoresis, Two-Dimensional
KW - Molecular Weight
KW - Oligopeptides
KW - Serine Proteinase Inhibitors
KW - Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
KW - Tight Junctions
M3 - Journal article
C2 - 10931115
SN - 0954-7894
VL - 30
SP - 1085
EP - 1096
JO - Clinical & Experimental Allergy
JF - Clinical & Experimental Allergy
IS - 8
ER -