a recombinant functional variant of the olive pollen allergen ole e 10 expressed in baculovirus system：一个橄榄花粉过敏原好啊e 10杆状病毒表达系统表达重组的功能变体.pdf

Journal of Biotechnology 121 (2006) 402–409 A recombinant functional variant of the olive pollen allergen Ole e 10 expressed in baculovirus system ´ ´ Patricia Barral, Alicia G. Serrano, Eva Batanero, Jesus Perez-Gil, ´ ´ ∗ Mayte Villalba, Rosalıa Rodrıguez Departamento de Bioqu´ımica y Biolog´ıa Molecular I, Facultad de Qu´ımicas, Universidad Complutense, 28040 Madrid, Spain Received 1 February 2005; received in revised form 8 July 2005; accepted 14 July 2005 Abstract Pollens have been reported as important sources of antigens causing type-I allergy and, among them, olive pollen has high clinical relevance in Mediterranean countries. The most recently described olive allergen, Ole e 10, is involved in cross-reactivity phenomena and related to asthma induction in allergic patients. These immunologic features make this allergen a good candidate to be included in diagnosis and therapy of protocols of allergic diseases. Since the availability of Ole e 10 from the olive pollen is limited, the allergen has been efficiently expressed in the baculovirus/insect cell system. The Ole e 10-cDNA inserted into the transfer vector pBacPAK8 allowed the expression of the recombinant protein in cultured Sf21 cells. Recombinant Ole e 10 (rOle e 10) was purified from the culture after dialysis and three chromatographic steps. Mass spectrometry, Edman degradation, IgE- and IgG-binding analyses were employed to characterize the recombinant allergen, which showed molecular and immunological equivalence with the natural protein. Affinity gel electrophoresis in presence of laminarin (1,3--gluc