The monoclonal antibody HNK-1 originally raised to an antigenic marker of natural killer cells also binds to selected regions in nervous tissue. The antigen is a carbohydrate that has attracted much interest as its expression is developmentally regulated in nervous tissue, and it is found, and proposed to be a ligand, on several of the adhesive glycoproteins of the nervous system. It is also expressed on glycolipids and proteoglycans, and is the target of monoclonal auto-antibodies that give rise to a demyelinating disease. The epitope, as characterized on glycolipids isolated from the nervous system, is expressed on 3-sulfated glucuronic acid joined by β1-3-linkage to a neolacto backbone. Here we exploit the neoglycolipid technology, in conjunction with immunodetection and in situ liquid secondary ion mass spectrometry, to characterize HNK-1-positive oligosaccharide chains derived by reductive alkaline release from total brain glycopeptides. The immunoreactive oligosaccharides detected are tetra- to octasaccharides that are very minor components among a heterogeneous population, each representing less than 0.1% of the starting material. Their peripheral and backbone sequences resemble those of the HNK-1-positive glycolipids. An unexpected finding is that they terminate not with N-acetylgalactosaminitol but with hexitol (2-substituted and 2,6-disubstituted). In a tetrasaccharide investigated in the greatest detail, the hexitol is identified as 2-substituted mannitol.

Sulfated galactan κ‐carrageenan is a linear polysaccharide with a repeating disaccharide sequence of alternating 4‐sulfated 3‐linked galactose and 4‐linked 3,6‐anhydrogalactose units. In contrast to many examples of chemical hydrolysis of polysaccharides, mild acid treatment of κ‐carrageenan resulted in facile and highly specific cleavage. In this article, we report the identification, by various MS and chromatographic techniques, of an unexpected series of exclusively odd‐numbered oligosaccharide fragments from its hydrolytic products. Detailed sequence analysis of the products indicated that all the oligosaccharide fragments have the 4‐sulfated 3‐linked galactose residues at both the reducing and the nonreducing ends. Further detailed investigation and analysis suggested that these odd‐numbered oligosaccharides were derived from two‐step cleavages of the glycosidic bonds on either sides of the 3,6‐anhydrogalactose residues. Neutral galactan agarose also contains 3,6‐anhydrogalactose and has a similar backbone sequence, and exhibited similar results upon mild acid hydrolysis. It is highly unusual to obtain exclusively odd‐numbered oligosaccharides from polysaccharides composed of ordered disaccharide repeats.

Human adenovirus 52 (HAdV-52) is one of only three known HAdVs equipped with both a long and a short fiber protein. While the long fiber binds to the coxsackie and adenovirus receptor, the function of the short fiber in the virus life cycle is poorly understood. Here, we show, by glycan microarray analysis and cellular studies, that the short fiber knob (SFK) of HAdV-52 recognizes long chains of α-2,8-linked polysialic acid (polySia), a large posttranslational modification of selected carrier proteins, and that HAdV-52 can use polySia as a receptor on target cells. X-ray crystallography, NMR, molecular dynamics simulation, and structure-guided mutagenesis of the SFK reveal that the nonreducing, terminal sialic acid of polySia engages the protein with direct contacts, and that specificity for polySia is achieved through subtle, transient electrostatic interactions with additional sialic acid residues. In this study, we present a previously unrecognized role for polySia as a cellular receptor for a human viral pathogen. Our detailed analysis of the determinants of specificity for this interaction has general implications for protein–carbohydrate interactions, particularly concerning highly charged glycan structures, and provides interesting dimensions on the biology and evolution of members of Human mastadenovirus G. Significance We present here that adenovirus type 52 (HAdV-52) attaches to target cells through a mechanism not previously observed in other human pathogenic viruses. The interaction involves unusual, transient, electrostatic interactions between the short fiber capsid protein and polysialic acid (polySia)-containing receptors on target cells. Knowledge about the binding interactions between polySia and its natural ligands is relatively limited, and our results therefore provide additional insight not only into adenovirus biology but also into the structural basis of polySia function. Since polySia can be found in high expression levels in brain and lung cancers where its presence is associated with poor prognosis, we suggest that this polySia-binding adenovirus could be useful for design of vectors for gene therapy of these cancers.