TY - JOUR
T1 - Molecular cloning of the mountain cedar (Juniperus ashei) pollen major allergen, Jun a 1
AU - Midoro-Horiuti, Terumi
AU - Goldblum, Randall M.
AU - Kurosky, Alexander
AU - Wood, Thomas G.
AU - Schein, Catherine H.
AU - Brooks, Edward G.
N1 - Copyright:
Copyright 2020 Elsevier B.V., All rights reserved.
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 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.
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 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.
KW - Allergen
KW - Cha o 1
KW - Chamaecyparis obtusa
KW - Cry j 1
KW - Cryptomeria japonica
KW - Jun a l
KW - Juniperus ashei
KW - Juniperus sabinoides
KW - Mountain cedar
KW - Pollinosis
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U2 - 10.1016/s0091-6749(99)70332-5
DO - 10.1016/s0091-6749(99)70332-5
M3 - Article
C2 - 10482836
AN - SCOPUS:0032825030
SN - 0091-6749
VL - 104
SP - 613
EP - 617
JO - Journal of Allergy and Clinical Immunology
JF - Journal of Allergy and Clinical Immunology
IS - 3 II
ER -