Characterization and gene cloning of an acidic thaumatin-like protein (TLP 1), an allergen from sapodilla fruit (Manilkara zapota).
BACKGROUND
Allergy to sapodilla (Manilkara zapota) fruit ingestion is rare. An independent study from our group has identified a basic thaumatin-like protein (TLP 2) as the major allergen. The present study was aimed at identifying and characterizing additional allergens from sapodilla.


METHODS
Allergic subjects were identified by case history, skin prick test (SPT) and allergen-specific IgE. Sapodilla extract was fractionated using SP-Sepharose into 3 components (SP1, SP2 and SP3) which were analyzed by native/SDS-PAGE, IgE-immunoblot, isoelectric focusing (IEF) and N-terminal sequencing. The conserved regions of plant TLPs and the N-terminal sequence were used to design primers for PCR.


RESULTS
SPT and ELISA confirmed a subject with oral allergy syndrome (OAS) to sapodilla and custard apple. Two proteins (26.9 and 24.5kDa; reducing conditions) were detected as allergens, of which the latter in SP2 has already been identified as basic TLP (TLP 2). The 26.9kDa protein present in SP1 was identified as an acidic TLP based on native PAGE, IEF and N-terminal sequencing. Presence of a basic β-1,3-glucanase in SP3 was inferred by zymography. Sequence analysis of the genomic clone of the acidic TLP gene revealed that it is intronless and non-glycosylated. Evolutionary relatedness to olive, grape and kiwi fruit allergenic TLPs were inferred by phylogenetic analysis.


CONCLUSIONS
An acidic TLP (TLP 1) was identified as a new allergen in sapodilla. TLP 1 is a single polypeptide (207 residues) belonging to the thaumatin family of the GH64-TLP-SF superfamily. Clinically, sapodilla should be considered in the list of fruits causing OAS.
Department of Biochemistry and Nutrition, Central Food Technological Research Institute (CFTRI), Council of Scientific and Industrial Research, Mysore, Karnataka, India.
Hassan G
HG
Ashok Kumar
Venkatesh L
VL
Hegde
Santoshkumar M
SM
Shetty
Yeldur P
YP
Venkatesh
D000293:Adolescent; D000328:Adult; D000485:Allergens / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D000595:Amino Acid Sequence; D052179:Antigens, Plant / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D005075:Biological Evolution; D003001:Cloning, Molecular; D005260:Female; D005512:Food Hypersensitivity / Q000175:diagnosis / Q000276:immunology*; D005638:Fruit / Q000276:immunology*; D006801:Humans; D007073:Immunoglobulin E / Q000097:blood; D008297:Male; D032129:Manilkara / Q000276:immunology*; D008875:Middle Aged; D008969:Molecular Sequence Data; D010802:Phylogeny; D010940:Plant Proteins / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D017422:Sequence Analysis, DNA; D012882:Skin Tests; D055815:Young Adult
D000485:Allergens; D052179:Antigens, Plant; D010940:Plant Proteins; C000589845:thaumatin-like protein 1, Manilkara zapota; D007073:Immunoglobulin E