Oligosaccharides on cathepsin D from porcine spleen

Cathepsin D from porcine spleen contained mannose (3.3%), glucosamine (1.4%), and mannose 6-phosphate (0.08%). Essentially all of the oligosaccharides of cathepsin D could be released by endo-β-N-acetylglucosaminidase H, pointing to oligomajmoside types of structures. Three neutral oligosaccharide fractions, containing 5, 6, and 7 mannose residues, respectively, were isolated by gel permeation chromatography on Bio-Gel P-2. Studies using exoglycosidase digestions and 500-MHz ¹H NMR spectroscopy revealed that their structures are [Manα1 → 2]_{0 or 1}Manα1 → 6[Manα1 → 3]Manα1 → 6[(Manα1 → 2)_{0 or 1}Manα1 → 3]Manβ1 → 4GlcNAcβ1 → 4 GlcNAc. These structures are identical to what have recently been proposed by Takahashi et al. for the major oligosaccharide units of cathepsin D from the same source (T. Takahashi P.G. Schimidt, and J. Tang (1983)J. Biol. Chem.258, 2819–2930), except for the occurrence of two isomeric oligosaccharides containing six mannoses. Only a part (3.4%) of the oligosaccharides were acidic, containing phosphates in monoester linkage. The phosphorylated oligosaccharides also consisted of oligomannoside-type chains which were analogous to, but more heterogeneous in size than the neutral oligosaccharides. Cathepsin D was bound to a mannose- and N-acetylglucosamine-specific lectin (mannan-binding protein) isolated from rabbit liver with the K_i value of 5.4 × 10^?6m.     

Die V?gel der Kurilen

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Die Lebensweise einiger wenig bekannter Sylviiden aus Ostasien

Ohne Zusammenfassung     

Serum lectin with known structure activates complement through the classical pathway

Serum mannan-binding protein (MBP), a lectin specific for mannose and N-acetylglucosamine, was revealed to activate the complement system as measured by passive hemolysis using sheep erythrocytes coated with yeast mannan. In contrast, rat liver MBP, which shares many properties in common with serum MBP, could not activate complement at all. The activation by serum MBP was inhibited effectively by the presence of haptenic sugars and dependent absolutely upon the presence of C4, indicating that the activation is initiated by the sugar binding activity of MBP and proceeds through the classical pathway. The 25 NH2-terminal amino acid sequence of rat serum MBP determined in this study was completely matched with that of MBP-A deduced from cDNA sequence by Drickamer et al. (Drickamer, K., Dordal, M. S., and Reynolds, L. (1986) J. Biol. Chem. 261, 6878-6887), revealing that MBP-A is in fact identical with serum MBP. On the basis of the knowledge of primary structures and physicochemical properties of rat serum and liver MBPs, a possible mechanism of the complement activation by serum MBP is discussed with reference to close similarity in the gross structures of serum MBP and C1q.     

Structures of oligosaccharides on beta-galactosidase from Aspergillus oryzae

The carbohydrate portions of beta-galactosidase from Aspergillus oryzae were found to be composed of two types of sugar chains. They were released equally well with endo-beta-N-acetylglucosaminidase H, but were distinct in their chain length. The long sugar chains (fraction I), corresponding to 4% of the total carbohydrate chains, were composed of galactomannan-type oligosaccharides, which consisted of mannose, galactose, glucose, and glucosamine in the molar ratios of 30.0, 16.4, 1.4, and 2.1 per mol of aspartic acid, respectively. The short sugar chains (fraction II), corresponding to 96% of the total carbohydrate chains, consisted of mannose, galactose, glucose, and glucosamine in the molar ratios of 9.4, 0.6, 0.3, and 1.7 per mol of aspartic acid, respectively. Both types of sugar chains were fractionated into neutral and acidic subfractions. The neutral subfraction of fraction I (I-N), corresponding to 1% of the total carbohydrate chains, was very heterogeneous in length and was resistant to digestion with alpha-mannosidase and beta-galactosidase. The neutral subfraction of fraction II (II-N), corresponding to 91% of the total carbohydrate, was composed of a mixture of oligosaccharides with oligomanneoside chains (Mann GlcNAcol). The major components were similar to high mannose-type oligosaccharides of mammalian origin in their composition and size (n = 5-9). However, digestion of II-N with alpha 1,2-mannosidase produced considerable amounts of Man6GlcNAcol, an unusual product in the case of high mannose-type oligosaccharides of mammalian origin, in addition to the common one, Man5GlcNAcol.     

Primary structure of rat liver mannan-binding protein deduced from its cDNA sequence

cDNA clones encoding rat liver mannan-binding protein (MBP), a lectin specific for mannose and N-acetylglucosamine, were isolated from a rat liver cDNA library carried in lambda gt 11, by screening with affinity purified polyclonal rabbit anti-rat liver MBP antibodies. The nucleotide sequence of the cDNA determined by the dideoxy method revealed the complete amino acid sequence of the MBP (226 residues). The NH2-terminal residue of the MBP, glutamic acid, was preceded by a predominantly hydrophobic stretch of 18 amino acids, which was assumed to be a signal peptide. Near the NH2-terminal, there was a collagen-like domain, which consisted of 19 repeats of the sequence Gly-X-Y. Here, X and Y were frequently proline and lysine. Three proline and lysine residues were hydroxylated, and one of the latter appeared to link to galactose. Computer analysis of several lectins for sequence homology suggested that the COOH-terminal quarter of the MBP is associated with the calcium binding as well as carbohydrate recognition.