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РЖ ВИНИТИ 34 (BI39) 96.04-04К1.13

Автор(ы) : Ellgaard, Lars, Holtet Thor L., Moestrup Soren K., Etzerodt, Mihcael, Thogersen Hans C.
Заглавие : Nested sts of protein fragments and their use in epitope mapping: Characterization of the epitope for the S4D5 monoclonal antibody binding to receptor associated protein
Источник статьи : J. Immunol. Meth. - 1995. - Vol. 180, N 1. - С. 53-61
Аннотация: The present report describes a new general procedure by which linear and some structure-dependent epitopes may be mapped in a protein antigen using a nested set of protein fragmets prepared from partial proteolysis products of a recombinant protein. The antigen, fused to an affinity tag, is partially fragmented and affinity sorted under denaturing conditions to produce a nested set of polypeptides, consisting of N- (or C-)terminal fragments. Immunoblots of SDS-PAGE fractionated sets of fragments are therefore directly readable in terms of molecular mass - i.e., approximate sequence positions - that identify sequence segments hardouring an epitope and any additional structural elements, required to maintain epitope conformation. Blots of N- and C-terminal nested sets of polypeptide fragments representing the human receptor associated protein (RAP) were prepared and probed with mAb S4D5. Fragments 1-177 and 94-323 were the shortest fragments detected by the antibody, suggesting the presence of an epitope within the 94-177 segment. Independent mapping based on recombinant fragments of the RAP homologue, rat Heymann nephritis antigen, confirmed that the epitope resides in the Pro[115]-Asp[177] segment. The model study demonstrates the utility of nested sets of protein fragments as fast and inexpensive tools for epitope mapping. Дания, Lab.Gene Expression, Dep. Chem., Univ. Aarhus; DK-8000 Aarhus
ГРНТИ : 34.43.05
Предметные рубрики: АНТИГЕНЫ
БЕЛКОВЫЕ
ЭПИТОПЫ
КАРТИРОВАНИЕ
МЕТОД
ФРАГМЕНТЫ ПРОТЕОЛИЗА
ГНЕЗДОВЫЕ СОЕДИНЕНИЯ
ИСПОЛЬЗОВАНИЕ
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