PURIFICATION OFMURINE HEAT SHOCK PROTEIN90-BETA (84 KDA) FROMMICE SPLEEN

	PURIFICATION OFMURINE HEAT SHOCK PROTEIN90-BETA (84 KDA) FROMMICE SPLEEN
Iraqi Journal of Biotechnology
Article 1, Volume 9, Issue 4, 2010, Pages 782-796
Abstract
ABSTRACTMurine heat shock protein 90 beta isotype (84kDa) was purified from thecytoplasm of BALB/c mice spleen cells. Since this protein dose not have an in vitroactivity assay and the presence of protein with molecular weight equal or near 84 kDawas followed in each purification step. Cytoplasm fractions of spleen cells containedproteins of molecular weight (MW) ranged from 86.0 to 20.9kDa as determined withSDS-PAGE analysis. The ammonium sulfate 70% saturation precipitate of cytoplasmfraction exhibited protein bands with molecular weight ranged from 40.7 to 86.1kDa.Ion exchange chromatography with DEAE-Cellulose column for the ammoniumsulfate precipitated cytoplasm proteins produced one prominent peak, eluted between0.5 and 0.7M NaCl. This peak contain four proteins with MW ranged from 19 to83.1kDa. During gel filtration through Sepharose 6B column for the peak eluted fromDEAE-cellulose column, a peak with estimated molecular weight of 87kDa wasselected among others produced. Second round of gel filtration through Sephacryl S-200 was conducted for the 87kDa selected fractions eluted from the previous gelfiltration step. The presence of one peak was emphasized and its estimated MW was83.176 kDa. The molecular weight and purity of this protein was checked again withSDS-PAGE 10%. The identity of the purified proteins as Hsp90β was confirmed withimmunofixation assay by gel electrophoresis on 1%agarose using standard.Key words: Heat shock, Protein 90, Spleen cells.
Keywords
Heat Shock; protein; Spleen cells

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