A novel recognition of the major allergen, Hum j 1, from

Background: Methods: During our work in isolating potential allergens from HJ pollen, we found a protein of around 10 kDa with obvious immunoglobulin E (IgE) binding activity than other fractions. This protein was then further identified by mass spectrometry, and its gene sequence was matched from our in-house built transcripts of HJ pollen based on the identified peptides. The gene was further confirmed by polymerase chain reaction (PCR) cloning, and the recombinant protein was prepared by Results: The 10 kDa protein with obvious IgE binding activity was finally purified from HJ pollen with a purity of 98%. The liquid chromatography-tandem mass spectrometry (LC-MS/MS) results identified 6 internal peptides, which covered 98.9% of the mature sequence of the 10 kDa protein with the N-terminal segment as DNCFENGMK. We surprisingly found that the 10 kDa protein was previously underrecognized Hum j 1 based on its limited sequence of only N-terminus. The sequence failed to be classified into any protein families by InterPro functional analysis, indicating the Hum j 1 could be the orphan protein. Both natural and recombinant Hum j 1 exhibited similar CD spectrum. The specific IgE binding rate against them was determined as 79% (11/14) in patients' serum with HJ pollen allergy by ELISA with a strong correlation in absorbance values (R Conclusion: These findings first provided solid evidence about the major allergen Hum j 1 in HJ pollen and resolved its unclear status. The defined sequence could enhance understanding of Hum j 1 properties and aid in producing high-quality recombinant allergens for diagnosing and treating HJ pollen allergy.