ISSN:
1398-9995
Source:
Blackwell Publishing Journal Backfiles 1879-2005
Topics:
Medicine
Notes:
Background: A rapid method for the purification of the major 43-kDa all_ergen of Cupressus arizonica pollen, Cup a 1, was developed. Methods: The salient feature was a wash of the pollen in acidic buffer, followed by an extraction of the proteins and their purification by chromatography. Immunoblotting, ELISA, and lectin binding were tested on both the crude extract and the purified Cup a 1. Biochemical analyses were performed to assess the Cup a 1 isoelectric point, its partial amino-acid sequence, and its glycan composition. Results: Immunochemical analysis of Cup a 1 confirmed that the all_ergenic reactivity is maintained after the purification process. Partial amino-acid sequencing indicated a high degree of homology between Cup a 1 and all_ergenic proteins from the Cupressaceae and Taxodiaceae families displaying a similar molecular mass. The purified protein shows one band with an isoelectric point of 5.2. Nineteen out of 33 sera (57%) from patients all_ergic to cypress demonstrated significant reactivity to purified Cup a 1. MALDI-TOF mass spectrometry indicated the presence of three N-linked oligosaccharide structures: GnGnXF3 (i.e., a horseradish peroxidase-type oligosaccharide substituted with two nonreducing N-acetylglucosamine residues), GGnXF3/GnGXF3 (i.e., GnGnXF with one nonreducing galactose residue), and (GF)GnXF3/Gn(GF)XF3 (with a Lewisa epitope on one arm) in the molar ratio 67:8:23. Conclusions: The rapid purification process of Cup a 1 all_owed some fine studies on its properties and structure, as well as the evaluation of its IgE reactivity in native conditions. The similarities of amino-acid sequences and some complex glycan stuctures could explain the high degree of cross-reactivity among the Cupressaceae and Taxodiaceae families.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1034/j.1398-9995.2001.103125.x
