Molecular cloning of the mountain cedar (Juniperus ashei) pollen major allergen, Jun a 1

Terumi Midoro-Horiuti, Randall M. Goldblum, Alexander Kurosky, Thomas G. Wood, Catherine H. Schein, Edward G. Brooks

Research output: Contribution to journalArticle

56 Scopus citations

Abstract

Background: Cedar pollens cause allergic disease in diverse geographic areas. We have recently purified and characterized the major mountain cedar (Juniperus ashei) pollen allergen, Jun a 1. Objective: A full-length complementary DNA for Jun a 1 was cloned and sequenced, and the recombinant protein was expressed. Methods: Messenger RNA from mountain cedar pollen was purified and Jun a 1 sequences were established with use of reverse transcriptase-PCR and primers based on the N-terminal amino acid sequence of Jun a 1 and the homologous protein Cry j 1. Portions of the nucleotide sequence were confirmed by comparison with N-terminal amino acid sequencing of the intact tryptic fragments of the purified native protein. Recombinant Jun a 1 was cloned into pET 30, expressed in BL21, and purified by HPLC, and its allergenicity was analyzed by Western blotting with patient sera. Results: Jun a 1 possesses a high level of amino acid sequence homology with Cha o 1 and Cry j 1, the major allergens of Japanese cypress and Japanese cedar. The amino acid sequence of a region with putative pectate lyase activity was identical to that of Cry j 1 and Cha o 1. Jun a 1 contained 2 potential N-glycosylation sites that were distinct from those found in Cry j 1. The IgE from patient sera bound recombinant Jun a 1 in Western blot analysis. Conclusion: The high degree of homology of Jun a 1 with Cha o 1 and Cry j 1 may explain the cross-reactivity of conifer pollens. Differences in N-glycosylation suggest little overlap of glycopeptide epitopes.

Original languageEnglish (US)
Pages (from-to)613-617
Number of pages5
JournalJournal of Allergy and Clinical Immunology
Volume104
Issue number3 II
StatePublished - Dec 1 1999

Keywords

  • Allergen
  • Cha o 1
  • Chamaecyparis obtusa
  • Cry j 1
  • Cryptomeria japonica
  • Jun a l
  • Juniperus ashei
  • Juniperus sabinoides
  • Mountain cedar
  • Pollinosis

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology

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    Midoro-Horiuti, T., Goldblum, R. M., Kurosky, A., Wood, T. G., Schein, C. H., & Brooks, E. G. (1999). Molecular cloning of the mountain cedar (Juniperus ashei) pollen major allergen, Jun a 1. Journal of Allergy and Clinical Immunology, 104(3 II), 613-617.