Membrane-bound heparin binding proteins from HL-60 cells purified in a two-step affinity chromatography differentially eluted with divalent cations.
Solubilized membrane proteins from HL-60 cells were separated by two-step affinity chromatography. Proteins eluted with MgCl2 in the first heparin-gel were applied to the second heparin-gel and eluted with CaCl2. The eluted proteins were analysed and purified by electrophoresis. N-terminal amino acid sequences of eight proteins on the characteristic bands were determined. Homology search for the sequences indicated that three microsomal proteins, two nuclear proteins and a glycolytic enzyme were eluted with divalent cations, whereas a nuclear ribonucleoprotein and a membrane-cytoskelton linker protein were not dissociated with divalent cations, but with 2 M NaCl. Heparin affinity chromatography combined with differential elution with divalent cations can be a useful method for separation of membrane proteins.
Imai
Katsuyuki
K
Department of Science of Human Life, City College of Mie, Issinden-Nakano, Tsu-shi, Mie 514-0112, Japan. imai_ katsu@rio.odn.ne.jp
Iida
Tsukimi
T
Takano
Yasuo
Y
Uozumi
Nobuyuki
N
D000595:Amino Acid Sequence; D023181:Antimicrobial Cationic Peptides; D001798:Blood Proteins / Q000737:chemistry / Q000302:isolation & purification*; D002352:Carrier Proteins / Q000737:chemistry / Q000302:isolation & purification*; D002413:Cations, Divalent; D002846:Chromatography, Affinity / Q000379:methods*; D004591:Electrophoresis, Polyacrylamide Gel; D018922:HL-60 Cells; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry / Q000302:isolation & purification*
C062991:AZU1 protein, human; D023181:Antimicrobial Cationic Peptides; D001798:Blood Proteins; D002352:Carrier Proteins; D002413:Cations, Divalent; D008565:Membrane Proteins