Immunomagnetic capture of lens membrane fractions containing steroid binding protein.
This study describes the use of magnetic Dynabeads to purify microsomes from a crude microsomal fraction. A 28 kDa membrane-associated protein is proposed to mediate the binding of progesterone and other steroid hormones to ocular lens membranes and the rapid-nongenomic actions of these steroids. The subcellular location of this membrane steroid binding protein (MSBP) was probed by capture of organelles containing MSBP by magnetic beads displaying an antibody to a cytoplasmic domain of the protein. The beads were exposed to a crude microsomal fraction from lens epithelia. Western blotting was used to identify captured organelles and confirm the presence of MSBP. Microsomes and trace fiber cell plasma membrane were captured. Microsomes contained the 28 kDa MSBP. Lens fiber cell membrane contained a 55 kDa immunoreactive protein. The role of this serendipitously recognized protein in binding of steroids is unknown.
Department of Biochemistry, Kirksville College of Osteopathic Medicine, 800 West Jefferson, Kirksville, MO 63501, USA.
Patricia S
PS
Sexton
Richard J
RJ
Cenedella
D000818:Animals; D015153:Blotting, Western; D002417:Cattle; D018189:Immunomagnetic Separation / Q000379:methods*; D007700:Kinetics; D007908:Lens, Crystalline / Q000166:cytology* / Q000378:metabolism; D008565:Membrane Proteins / Q000302:isolation & purification*; D008861:Microsomes / Q000378:metabolism*; D017434:Protein Structure, Tertiary
D008565:Membrane Proteins; C433704:membrane steroid binding protein, lens