Purification of the Synaptic Membrane Glycoprotein D2 from Rat Brain

Abstract

D2 is a nervous‐specific membrane protein enriched in fractions of synaptosomal membranes from rat brain. Recently, an immunochemical relationship between D2 and the chick cell adhesion molecule (CAM) has been demonstrated. There is reason to believe thai D2 is involved in adhesion phenomena between neurites. The purpose of the present study was to purify and further characterize the D2 protein from rat brain. In the developed purification procedure synaptosomal membranes from rat brains were prepared and solubilized by means of non‐ionic detergent. The subsequent purification steps were hyroxylapatite chromatography, wheat germ lcctin affinity chromatography, gel filtration, and lysine affinity chromatography. The purified D2 was found to be enriched 240 times compared with the starting brain homogenate and 120 times compared with the synaptosomal membrane fraction. The recovery of D2 was 26% when the amount of D2 in ihe synaptosomal membrane fraction was set to 100%. The purified D2 antigen was used for production of monospecific rabbit anlisera, and it was found to be composed of two polypeptides of apparent molecular weights 130,000 and 150,000, respectively.