Abstract. Recently, poly α2,8 deaminoneuraminic acid (poly α2,8 KDN) was demonstrated in various embryonic and adult mammalian tissues. This study reports the purification and characterization of the single poly α2,8 KDN-bearing glycoprotein from rat kidney. Amino acid sequences of proteolytic fragments shared homology with megalin, a member of the LDL receptor family. Immunochemical analysis supported this finding, since immunoprecipitated poly α2,8 KDN-bearing glycoprotein was immunoreactive with anti-megalin antibodies in Western blotting and conversely immunoprecipitated megalin was immunoreactive with the monoclonal anti-poly α2,8 KDN antibody. Furthermore, receptor-associated protein affinity-purified megalin reacted with the anti-poly α2,8 KDN antibody. By immunoelectron microscopy, labeling for both poly α2,8 KDN and megalin coincided in the brush border, endocytic invaginations and vesicles, and apical dense tubules of proximal convoluted tubules. Immunoreactivity for poly α2,8 KDN on purified megalin was abolished by β-elimination reaction but not by N-glycosidase F treatment. These data identified megalin as the sole glycoprotein of rat kidney, which contains poly α2,8 KDN present on O-glycosidically linked oligosaccharides. Furthermore, this study shows that megalin carries N-glycosidically linked hybrid and complex-type oligosaccharides terminating with sialic acid. Both poly α2,8 KDN and sialic acids on megalin may contribute to the binding of Ca2+ and cationic ligands.
Characterization of the spectrum of alternative splicing of alpha 1 (XIII) collagen transcripts in HT-1080 cells and calvarial tissue resulted in identification of two previously unidentified alternatively spliced sequences, one previously unidentified exon, and nine new mRNA variants.