Iida Tsukimi, Kamo Masaharu, Uozumi Nobuyuki, Inui Takashi, Imai Katsuyuki
Department of Science of Human Life, City College of Mie, 157, Ishinnden-nakano, Tsu, Mie 514-0112, Japan.
J Chromatogr B Analyt Technol Biomed Life Sci. 2005 Sep 5;823(2):209-12. doi: 10.1016/j.jchromb.2005.06.025.
Membrane proteins were obtained from the mitochondrial fraction of HL-60 cells by solubilization with octyl glucoside and bound to heparin-gels. Bound proteins were successively eluted with solutions containing increasing concentrations of Mg(2+) in the first and increasing concentrations of Ca(2+) in the second chromatography. After SDS-PAGE and subsequent N-terminal amino acid analysis of proteins on each band, 13 proteins were identified. Fifteen out of the 37 proteins analysed were modified at their N-termini. These results show that this two-step affinity chromatography method using divalent cations as eluents can be applied to a variety of membranes for the isolation of specific proteins.
膜蛋白通过用辛基葡糖苷溶解从HL-60细胞的线粒体部分获得，并与肝素凝胶结合。结合的蛋白在第一次色谱中用含有浓度递增的Mg(2+)的溶液依次洗脱，在第二次色谱中用含有浓度递增的Ca(2+)的溶液洗脱。经过SDS-PAGE以及随后对每条带上的蛋白进行N端氨基酸分析，鉴定出了13种蛋白。在分析的37种蛋白中，有15种在其N端被修饰。这些结果表明，这种以二价阳离子作为洗脱剂的两步亲和色谱法可应用于多种膜以分离特定蛋白。
