ANALYSIS OF GLYCOPROTEINS IN THE GLOMERULAR BASEMENT MEMBRANE OF STREPTOZOTOCIN DIABETIC RATS : USE OF LECTINS FOR DETECTION OF ELECTROPHORETICALLY SEPARATED GLYCOPROTEINS

This study was performed to clarify the composition of glycoproteins in the glomerular basement membrane (GBM) isolated from streptozotocin (STZ) diabetic rats. Four kinds of lectin were used for detection of the glycoproteins which were electrophoretically separated from the purified GBM. STZ diabetic rats were induced by a single intravenous injection of STZ 65 mg/kg body weight. GBM was obtained by sonication of glomeruli isolated through the sieving method. Glycoproteins solubilized with 8M urea from the GBM were separated by polyacrylamide gel electrophoresis in sodium dodecyl sulfate and transferred onto nitrocellulose sheets and polyvinylidene difluoride (PVDF) membranes. The glycoproteins were then stained immunochemically with four lectins. Staining patterns of Ricinus communis agglutinin and Phaseolus vulgaris agglutinin of STZ diabetic rats were similar to those of normal rats. However, unlike in normal rats, in STZ diabetic rats one major band was identified by wheat germ agglutinin and Concanavalin A. This band was detected at an apparent molecular weight of 60,000. The amino acid composition of this glycoprotein, electroblotted onto PVDF membrane, was analysed by HPLC after hydrolysis. This glycoprotein did not contain hydroxyproline or hydroxylysine, which are abundant in the type Ⅳ collagen. Furthermore, the amino acid composition of this glycoprotein was different from that of known components of the GBM