Identification and cloning of prs a 1, a 32-kDa endochitinase and major allergen of avocado, and its expression in the yeast Pichia pastoris

J Biol Chem. 1998 Oct 23;273(43):28091-7. doi: 10.1074/jbc.273.43.28091.

Abstract

Avocado, the fruit of the tropical tree Persea americana, is a source of allergens that can elicit diverse IgE-mediated reactions including anaphylaxis in sensitized individuals. We characterized a 32-kDa major avocado allergen, Prs a 1, which is recognized by 15 out of 20 avocado- and/or latex-allergic patients. Natural Prs a 1 was purified, and its N-terminal and two tryptic peptide sequences were determined. We isolated the Prs a 1 encoding cDNA by PCR using degenerate primers and 5'-rapid amplification of cDNA ends. The Prs a 1 cDNA coded for an endochitinase of 326 amino acids with a leader peptide of 25 amino acids. We expressed Prs a 1 in the yeast Pichia pastoris at 50 mg/liter of culture medium. The recombinant Prs a 1 showed endochitinase activity, inhibited growth and branching of Fusarium oxysporum hyphae, and possessed IgE binding capacity. IgE cross-reactivity with latex proteins including a 20-kDa allergen, most likely prohevein, was demonstrated, providing an explanation for the commonly observed cross-sensitization between avocado and latex proteins. Sequence comparison showed that Prs a 1 and prohevein had 70% similarity in their chitin-binding domains. Characterization of chitinases as allergens has implications for engineering transgenic crops with increased levels of chitinases.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Allergens / biosynthesis
  • Allergens / genetics*
  • Allergens / isolation & purification
  • Amino Acid Sequence
  • Antigens, Plant
  • Base Sequence
  • Chitinases / biosynthesis
  • Chitinases / genetics*
  • Chitinases / isolation & purification
  • Cloning, Molecular
  • Cross Reactions
  • DNA, Complementary / genetics
  • Humans
  • Hypersensitivity
  • Immunoglobulin E / metabolism
  • Latex Hypersensitivity
  • Lauraceae / genetics*
  • Lauraceae / immunology
  • Molecular Sequence Data
  • Pichia / genetics
  • Plant Proteins / immunology
  • Recombinant Proteins / biosynthesis
  • Recombinant Proteins / isolation & purification
  • Sequence Analysis, DNA
  • Sequence Homology, Amino Acid
  • Substrate Specificity

Substances

  • Allergens
  • Antigens, Plant
  • DNA, Complementary
  • Plant Proteins
  • Recombinant Proteins
  • Immunoglobulin E
  • Chitinases
  • Prs a 1 allergen

Associated data

  • GENBANK/Z78202