Molecular cloning and expression in insect cells of honeybee venom allergen acid phosphatase (Api m 3).

Acid phosphatase (Api m 3) is a major allergen in honeybee (Apis mellifera) venom, and its availability as a recombinant protein may facilitate the development of improved diagnostic tests and immunotherapies.
One objective is the determination of the complete primary structure of Api m 3 and to obtain recombinant Api m 3 on the basis of expression in insect cells. Another objective is the quantitative analysis of patient serum IgE antibody reactive to recombinant Api m 3.
The cloning of Api m 3 from venom gland cDNA and its expression as a full-length protein in eukaryotic insect cells is described. The immunoreactivity of serum IgE antibodies of honeybee venom-sensitized patients to recombinant Api m 3 was determined in an enzyme immunoassay.
PCR amplification generated a 1122-bp DNA fragment whose identity as the coding sequence of Api m 3 was verified by several means. Recombinant Api m 3, expressed in Trichoplusia ni cells, showed an expected molecular weight and enzymatic activity at pH 4.5. Analysis of tryptic fragments of purified recombinant Api m 3 by mass spectrometry confirmed its identity. In immunoassays, recombinant Api m 3 is specifically recognized by IgE antibodies of pooled serum in Western blots and by 37% of the individual sera of honeybee venom-sensitized patients in ELISA analysis.
The availability of recombinant Api m 3 provides a tool for both the development of improved diagnostic tests and the design of safer and more effective immunotherapeutic approaches for honeybee venom allergy.
The recombinant venom allergen Api m 3 is a key element in the search for an optimized component-resolved approach to honeybee venom allergy with regard to both the development of superior diagnostic tests and the improvement of allergen immunotherapy.
AuthorsThomas Grunwald, Benjamin Bockisch, Edzard Spillner, Johannes Ring, Reinhard Bredehorst, Markus W Ollert
JournalThe Journal of allergy and clinical immunology (J Allergy Clin Immunol) Vol. 117 Issue 4 Pg. 848-54 (Apr 2006) ISSN: 0091-6749 [Print] United States
PMID16630944 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
Chemical References
  • Allergens
  • Bee Venoms
  • DNA, Complementary
  • Immunoglobulin E
  • Acid Phosphatase
  • Acid Phosphatase (genetics, immunology)
  • Allergens (genetics)
  • Amino Acid Sequence
  • Anaphylaxis (etiology, therapy)
  • Animals
  • Base Sequence
  • Bee Venoms (genetics, immunology)
  • Bees (enzymology, genetics, immunology)
  • Cell Line
  • Cloning, Molecular
  • DNA, Complementary (genetics)
  • Desensitization, Immunologic
  • Gene Expression
  • Genes, Insect
  • Humans
  • Immunoglobulin E (blood)
  • In Vitro Techniques
  • Insect Bites and Stings (complications, immunology)
  • Molecular Sequence Data
  • Moths
  • Sequence Homology, Amino Acid

Join CureHunter, for free Research Interface BASIC access!

Take advantage of free CureHunter research engine access to explore the best drug and treatment options for any disease. Find out why thousands of doctors, pharma researchers and patient activists around the world use CureHunter every day.
Realize the full power of the drug-disease research network!

Choose Username:
Verify Password: