Isolation of Influenza Virus A Hemagglutinin C-Terminal Domain by Hemagglutinin Proteolysis in Octylglucoside Micelles
Abstract:A method of isolation of hydrophobic membrane-bound C-terminal domain of influenza virus A hemagglutinin (HA) is suggested. The method is based on the insertion of HA into octylglucoside micelles followed by pepsin or thermolysin hydrolysis. Subsequent treatment of proteolytic digests with chloroform-hexafluoroisopropanol mixture resulted in the extraction of a few hydrophobic peptides into organic phase. Mass-spectrometry (MALDI-TOF) analysis revealed that the peptides with ion masses corresponding to the anchoring C-terminal domain with or without modifications predominated in the organic solution. The data obtained confirmed our speculation on the possibility of the suggested isolation scheme following from the strong interactions of anchoring domains in compact trimeric structure of HA spikes combined with micelle protection effect. Several appropriate peptides presence in the organic phase apparently arises from the presence of a few accessible proteolytic sites in HA transmembrane region.
Document Type: Research Article
Affiliations: Department of Chromatography, A.N. Belozersky Institute of Physico-Chemical Biology, Moscow State University, Leninskie Gory, Moscow 119992, Russia.
Publication date: September 1, 2006
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