Glycoproteins and proteoglycans of the chromaffin granule matrix.

Chromaffin granules have been purified from bovine adrenal medulla and the soluble proteins fractionated by chromatography on DEAE-cellulose. This procedure allowed isolation of the chromogranins (a class of related acidic glycoproteins) and of two proteoglycans, as well as a fraction highly enriched in dopamine 8hydroxylase. Oligosaccharides and glycopeptides were fractionated and purified by ion-exchange, thin layer, and affinity chromatography, and their structural properties were studied by methylation analysis and specific degradations. The chromogranins contain 5.4% carbohydrate consisting predominantly of 0-glycosidically linked triand tetrasaccharides composed of N-acetylgalactosamine, galactose, and sialic acid (N-acetyl-(AcNeu) and/ o r N-glycolylneuraminic (GcNeu) acid), as well as a small proportion of N-glycosidically linked oligosaccharide units. One-half of the 0-glycosidically linked carbohydrate is released by alkaline borohydride in the form of the trisaccharides AcNeu(a2 + 3)Gal(81 -P 3)GalNAcol (N-acetyl-D-galactosaminitol) (40%) and GcNeu(a2 + 3)Gal(81-+ 3)GalNAcol(lO%). The tetrasaccharide components consist of AcNeu(a2 + 3)Gal(fl1 + 3)[AcNeu(a2 + 6)IGalNAcol (20%), GcNeu(a2 -P 8)GcNeu(a2 + 6)[Gal(fll+ 3)]GalNAcol(7%), as well as 16 and 78 , respectively, of the tetrasaccharide structures given above but in which the two types of sialic acid cannot be individually assigned to specific linkage positions. After fractionation of the chromogranins by gel filtration, it was found that these oligosaccharides are not uniformly distributed as a function of chromogranin size, insofar as the largest chromogranins (chromogranin A) contain predominantly trisaccharides, whereas the tetrasaccharides are concentrated in the chromogranins of intermediate molecular size. The chromaffin granule matrix also contains two proteoglycans (I and 11) in which the glycosaminoglycan component consists of 48% dermatan sulfate, 2324% each of chondroitin 4and 6-sulfate, and 5% heparan sulfate. Nand 0-glycosidically linked glycoprotein oligosaccharides are also present. Although the two proteoglycans appear to have very similar o r identical protein moieties, proteoglycan 11 has twice the concentration of glycosaminoglycans and one-halfthe concentration of 0-glycosidically linked oligosaccharides (primarily disialyl derivatives of Gal(B1 + 3)GalNAc) as compared to proteoglycan I. .* This work was supported by Grants NS-09348 and NS-13876 from the National Institutes of Health, Grant BNS-7915410 from the National Science Foundation, and a grant from the Sigrid Juse1iu.q Foundation, Finland. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.