Identification of beta(2)-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. beta(2)-glycoprotein I was identified as a calmodulin-binding protein both by N-terminal sequencing and by immunoblotting. Quantification showed that beta(2)-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 beta(2)-glycoprotein I binds calmodulin and that beta(2)-glycoprotein I may in kidney be a membrane-associated protein. Immunohistochemical studies identified beta(2)-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 beta(2)-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 beta(2)-glycoprotein I being a marker for renal tubular disease.