Purification and structural analysis of the novel glycoprotein allergen Cyn d 24, a pathogenesis-related protein PR-1, from Bermuda grass pollen

FEBS J. 2005 Dec;272(24):6218-27. doi: 10.1111/j.1742-4658.2005.05000.x.

Abstract

Bermuda grass pollen (BGP) contains a very complex mixture of allergens, but only a few have been characterized. One of the allergens, with an apparent molecular mass of 21 kDa, has been shown to bind serum IgE from 29% of patients with BGP allergy. A combination of chromatographic techniques (ion exchange and reverse phase HPLC) was used to purify the 21 kDa allergen. Immunoblotting was performed to investigate its IgE binding and lectin-binding activities, and the Lysyl-C endopeptidase digested peptides were determined by N-terminal sequencing. The cDNA sequence was analyzed by RACE PCR-based cloning. The protein mass and the putative glycan structure were further elucidated using MALDI-TOF mass spectrometry. The purified 21 kDa allergen was designated Cyn d 24 according to the protocol of International Union of Immunological Societies (IUIS). It has a molecular mass of 18,411 Da by MALDI-TOF analysis and a pI of 5.9. The cDNA encoding Cyn d 24 was predicted to produce a 153 amino acid mature protein containing tow conserved sequences seen in the pathogen-related protein family. Carbohydrate analysis showed that the most abundant N-linked glycan is a alpha(3)-fucosylated pauci-mannose (Man3GlcNAc2) structure, without a Xyl beta-(1,2)-linked to the branching beta-Man. Thus, Cyn d 24 is a glycoprotein and the results of the sequence alignment indicate that this novel allergen is a pathogenesis-related protein 1. To the best of our knowledge, this is the first study to identify any grass pollen allergen as a pathogenesis-related protein 1.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Allergens / chemistry*
  • Allergens / isolation & purification
  • Amino Acid Sequence
  • Antigens, Plant / chemistry*
  • Antigens, Plant / isolation & purification
  • Carbohydrates / analysis
  • Chromatography
  • Cynodon / immunology*
  • Glycoproteins / chemistry*
  • Glycoproteins / isolation & purification
  • Immunoglobulin E / metabolism
  • Lectins / metabolism
  • Molecular Sequence Data
  • Plant Proteins / chemistry*
  • Plant Proteins / isolation & purification
  • Pollen / immunology*
  • Sequence Alignment

Substances

  • Allergens
  • Antigens, Plant
  • Carbohydrates
  • Cyn d 24 allergen, Cynodon dactylon
  • Glycoproteins
  • Lectins
  • Plant Proteins
  • pathogenesis-related proteins, plant
  • Immunoglobulin E