Isolation and biochemical characterization of a thaumatin-like kiwi allergen.
BACKGROUND
Kiwi fruit allergy, as well as its association with hypersensitivity to other foods and to pollen, has been extensively reported in the last few years. Several IgE-binding components have been detected in kiwi extract, but only one 30- kd allergen has been isolated; it was identified as actinidin (Act c 1). Recently, we have reported a 24-kd kiwi protein to be a potential major allergen in a group of patients with oral allergy syndrome (OAS).


OBJECTIVE
The aim of this study was to purify and characterize the 24-kd kiwi allergen biochemically.


METHODS
Seven polysensitized patients with OAS to kiwi were used in this study. The kiwi allergen was isolated by using a combination of gel permeation, ion exchange, and immobilized metal ion affinity chromatography. Its biochemical characterization included determination of its isoelectric point, molecular weight, N-terminal sequencing, concanavalin A -binding ability, digestibility in simulated gastric fluid, and antifungal activity. Western blotting, 2-dimensional PAGE immunoblotting, and skin prick tests were performed to characterize the isolated protein immunochemically.


RESULTS
All 7 patients recognized the isolated 24-kd kiwi protein as an allergen. The isolated protein consisted of 2 isoforms with isoelectric points of 9.4 and 9.5 migrated as one protein band of 20 kd after SDS-PAGE under nonreducing conditions or at 24 kd under reducing conditions. The partial N-terminal sequence revealed that it is a thaumatin-like protein (TLP) with concanavalin A -binding ability. The protein showed antifungal activity toward Saccharomyces carlsbergensis, and Candida albicans. The protein was degraded by the simulated gastric fluid within 1 minute. Both isoforms bound IgE from a pool of sera in a 2-dimensional PAGE immunoblot. The TLP elicited positive skin prick test responses in 4 (80 %) of 5 patients with OAS.


CONCLUSIONS
This study reported isolation and full characterization of a new kiwi allergen, TLP (isoelectric points of 9.4 and 9.5 and molecular weight of 24 kd), which belongs to the family of pathogenesis-related proteins. The isolated protein expressed antifungal activity toward S carlsbergensis and C albicans.
Gavrović-Jankulović
Marija
M
Department of Biochemistry, Faculty of Chemistry, University of Belgrade, Yugoslavia.
ćIrković
Tanja
T
Vucković
Olga
O
Atanasković-Marković
Marina
M
Petersen
Arnd
A
Gojgić
Gordana
G
Burazer
Lidija
L
Jankov
Ratko M
RM
D029042:Actinidia / Q000276:immunology*; D000485:Allergens / Q000737:chemistry* / Q000276:immunology / Q000302:isolation & purification*; D000595:Amino Acid Sequence; D000935:Antifungal Agents / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology; D005512:Food Hypersensitivity / Q000175:diagnosis / Q000276:immunology*; D005638:Fruit / Q000276:immunology; D006801:Humans; D007073:Immunoglobulin E / Q000276:immunology; D007526:Isoelectric Point; D008970:Molecular Weight; D010940:Plant Proteins / Q000737:chemistry / Q000276:immunology* / Q000302:isolation & purification; D013549:Sweetening Agents / Q000737:chemistry
D000485:Allergens; D000935:Antifungal Agents; D010940:Plant Proteins; D013549:Sweetening Agents; D007073:Immunoglobulin E; C003427:thaumatin protein, plant