Identification of beta2-glycoprotein I as a membrane-associated protein in kidney: purification by calmodulin affinity chromatography.
Outer renal medulla calmodulin-binding proteins from a soluble protein fraction and a plasma membrane fraction solubilized in CHAPS were retained on a calmodulin-Sepharose 4B column in the presence of Ca2+, and subsequently eluted by EGTA. The calmodulin-binding proteins constituted 2.5% of the soluble protein and 0.1% of the solubilized membrane protein. beta2-glycoprotein I was identified as a calmodulin-binding protein both by N-terminal sequencing and by immunoblotting. Quantification showed that beta2-glycoprotein I constituted the major part (approx. 35%) of the calmodulin-binding membrane proteins, but only a minor part (approx. 0.1%) of the calmodulin-binding proteins in the soluble fraction. These results show for the first time that beta2-glycoprotein I binds calmodulin and that beta2-glycoprotein I may in kidney be a membrane-associated protein. Immunohistochemical studies identified beta2-glycoprotein I in several parts of the cortex and the medulla of the kidney, including Bowman's capsula, the tubular lumen and the tubular epithelium, indicating that beta2-glycoprotein I, despite its relatively high molecular mass, is filtrated in the glomerulus and subsequently reabsorbed by the tubular epithelium. This is in agreement with beta2-glycoprotein I being a marker for renal tubular disease.
Klaerke
D A
DA
Biomembrane Research Center, Department of Medical Physiology, The Panum Institute, University of Copenhagen, Copenhagen N, Denmark.
Røjkjaer
R
R
Christensen
L
L
Schousboe
I
I
D000595:Amino Acid Sequence; D000818:Animals; D002148:Calmodulin-Binding Proteins / Q000378:metabolism*; D002846:Chromatography, Affinity; D006023:Glycoproteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D015151:Immunoblotting; D007150:Immunohistochemistry; D007679:Kidney Medulla / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D013552:Swine; D053482:beta 2-Glycoprotein I
D002148:Calmodulin-Binding Proteins; D006023:Glycoproteins; D008562:Membrane Glycoproteins; D053482:beta 2-Glycoprotein I