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

Terumi Midoro-Horiuti, R. M. Goldblum, A. Kurosky, Thomas Wood, C. H. Schein, E. G. Brooks

Research output: Contribution to journalArticle

56 Citations (Scopus)

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 I 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 I
DOIs
StatePublished - 1999

Fingerprint

Molecular Cloning
Pollen
Glycosylation
Amino Acid Sequence
Western Blotting
Cupressus
Cryptomeria
Coniferophyta
Amino Acid Sequence Homology
Glycopeptides
Protein Sequence Analysis
Reverse Transcriptase Polymerase Chain Reaction
Serum
Recombinant Proteins
Allergens
Immunoglobulin E
Epitopes
Proteins
Complementary DNA
High Pressure Liquid Chromatography

Keywords

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

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology

Cite this

Molecular cloning of the mountain cedar (Juniperus ashei) pollen major allergen, Jun a 1. / Midoro-Horiuti, Terumi; Goldblum, R. M.; Kurosky, A.; Wood, Thomas; Schein, C. H.; Brooks, E. G.

In: Journal of Allergy and Clinical Immunology, Vol. 104, No. 3 I, 1999, p. 613-617.

Research output: Contribution to journalArticle

Midoro-Horiuti, Terumi ; Goldblum, R. M. ; Kurosky, A. ; Wood, Thomas ; Schein, C. H. ; Brooks, E. G. / Molecular cloning of the mountain cedar (Juniperus ashei) pollen major allergen, Jun a 1. In: Journal of Allergy and Clinical Immunology. 1999 ; Vol. 104, No. 3 I. pp. 613-617.
@article{fe99873b2d2f4eaf95dc96e158610f89,
title = "Molecular cloning of the mountain cedar (Juniperus ashei) pollen major allergen, Jun a 1",
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 I 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.",
keywords = "Allergen, Cha o 1, Chamaecyparis obtusa, Cry j 1, Cryptomeria japonica, Jun a 1, Juniperus ashei, Juniperus sabinoides, Mountain cedar, Pollinosis",
author = "Terumi Midoro-Horiuti and Goldblum, {R. M.} and A. Kurosky and Thomas Wood and Schein, {C. H.} and Brooks, {E. G.}",
year = "1999",
doi = "10.1016/S0091-6749(99)70332-5",
language = "English (US)",
volume = "104",
pages = "613--617",
journal = "Journal of Allergy and Clinical Immunology",
issn = "0091-6749",
publisher = "Mosby Inc.",
number = "3 I",

}

TY - JOUR

T1 - Molecular cloning of the mountain cedar (Juniperus ashei) pollen major allergen, Jun a 1

AU - Midoro-Horiuti, Terumi

AU - Goldblum, R. M.

AU - Kurosky, A.

AU - Wood, Thomas

AU - Schein, C. H.

AU - Brooks, E. G.

PY - 1999

Y1 - 1999

N2 - 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 I 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.

AB - 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 I 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.

KW - Allergen

KW - Cha o 1

KW - Chamaecyparis obtusa

KW - Cry j 1

KW - Cryptomeria japonica

KW - Jun a 1

KW - Juniperus ashei

KW - Juniperus sabinoides

KW - Mountain cedar

KW - Pollinosis

UR - http://www.scopus.com/inward/record.url?scp=0032825030&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0032825030&partnerID=8YFLogxK

U2 - 10.1016/S0091-6749(99)70332-5

DO - 10.1016/S0091-6749(99)70332-5

M3 - Article

VL - 104

SP - 613

EP - 617

JO - Journal of Allergy and Clinical Immunology

JF - Journal of Allergy and Clinical Immunology

SN - 0091-6749

IS - 3 I

ER -