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J Allergy Clin Immunol. 1999 Sep;104(3 Pt 1):613-7.

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

Author information

1
Department of Pediatrics, Child Health Research Center, University of Texas Medical Branch at Galveston, 77555-0366, USA.

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.

PMID:
10482836
[Indexed for MEDLINE]

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