The artificial sulfated sialoglycopolypeptides (5 and 6) carrying the Neu5Acα2,3Galβ1,4 (6-sulfo)GlcNAc and Neu5Acα2,6Galβ1,4(6-sulfo)GlcNAc residues in the side chain were designed as hemagglutinin inhibitors against influenza viruses. 5-Trifluoroacetamidopentyl β-6-O-sulfo-N-acetylglucosaminide (5-TFAP-β-6SGN, 1) was first produced by β-N-acetylhexosaminidase-mediated transglycosylation using β-D-6-O-sulfo-GlcNAc-O-pNP (6SGN-β-pNP) as the donor and 5-trifluoroacetamido-1-pentanol as the acceptor. Galactosylation with glycosyltransferase was carried out to afford the key disaccharide, 6-O-sulfated N-acetyllactosaminide derivative 2. After detrifluoroacetylation of 2, 6-O-sulfated N-acetyllactosaminide 3 with the 5-aminopentyl group at the reducing end was obtained in three steps in 62% yield. 6-O-Sulfated disaccharide 3 was then coupled with the carboxyl groups of γ-polyglutamic acid (γ-PGA) by a conventional BOP/HOBt chemistry, giving glycopolypeptide 4. Trans-sialylation of glycopolypeptide 4 with α2,3-sialyltransferase or α2,6-sialyltransferase gave the corresponding sulfated sialoglycopolypeptides 5 and 6, respectively. The binding affinity of sulfated sialoglycopolypeptides (5 and 6) to influenza virus hemagglutinin was examined using a hemagglutination inhibition assay. The sulfated α2,6-sialoglycopolypeptide (6) selectively inhibited hemagglutination mediated by human virus A/Aichi/2/68 (H3N2) and had a relative binding affinity for hemagglutinin of ca. 4.9 × 102-fold higher than that of the naturally occurring fetuin control. The sulfated α2,3-sialoglycopolypeptide (5) selectively inhibited hemagglutination mediated by avian virus A/Duck/HongKong/313/4/78 (H5N3) and displayed an approximately 1.2 × 103-fold higher affinity over fetuin. In addition, the bindingaffinity of 5 was slightly higher than that of the non-sulfated α2,3 sialoglycopolypeptide (7).
The kinetics of sucrose hydrolysis was investigated in a water-ethanol mixture with ethanol concentrations of 0–80% (v/v) under subcritical conditions in the 160–190°C range. Sucrose underwent autocatalytic hydrolysis in the subcritical mixtures. The rate of sucrose hydrolysis to glucose and fructose was reduced with increasing ethanol concentration, and ethanol showed a dilution effect on the conversion. The temperature dependence of the reaction rate constant for sucrose hydrolysis obeyed the Arrhenius equation. The fructose and glucose products underwent further decomposition, and the yield of fructose was much lower than that of glucose when the ethanol concentration increased. Thus, ethanol exerted other effects on the reaction in the subcritical water-ethanol mixture.
Cassava starch granules were treated with three types of amylase and the resulting insoluble starch granules collected and their gelling properties investigated. Gel produced from starch granules treated with either α-amylase from Bacillus amyloliquefaciens or β-amylase from barley was slightly weaker than that from native cassava starch granules. On the other hand, starch granules treated with α-amylase from Aspergillus niger produced starch gel with significantly enhanced hardness and elasticity. This observation is surprising because very limited hydrolysis (2.8%) of starch produces such a great impact on gelling properties of starch and because it contradicts the generally held view that partially hydrolyzed starch shows decreased gel-forming ability. In order to understand the mechanism behind this, we analyzed the starch component leached from starch granules into hot water using gel permeation chromatography. The results suggest that enzymatic treatment of starch granules significantly changes the properties of starch by altering the composition of leached material (matrix) from swelled starch granules (filler).
D-mannose isomerase was cloned and characterized from a newly isolated actinobacteria strain, Thermobifida fusca MBL10003. The structural gene (manI) is predicted to encode a polypeptide of 407 amino acids with an estimated molecular mass of 43,900. Although the identity of the deduced amino acid sequence is not so high (45.7% to Salmonella enterica, and 38.5% to Agrobacterium radiobacter), the catalytic center appears has an essential structure that is conserved in the characterized homologs. It was a dimeric enzyme composed of two active monomer units. The optimal temperature and pH were 60°C and 8.0, respectively. The enzyme was stable up to around 60°C, and between pH 4 and 11. It showed activity on D-mannose and also on D-lyxose, but not on N-acetyl D-glucosamine, suggesting that it is functionally different from N-acyl D-glucosamine 2-epimerase despite the sequence similarity. Although Km value for D-mannose, 115 mM was similar to other mannose isomerase, kcat, hence kcat/Km was much higher than those. The enzyme was significantly inhibited by such divalent metal ions as Cu2+, Cd2+ or Ca2+, but it was not a metal-required enzyme for activity emergence.
The physicochemical properties of starch from Kaga lotus (Shina-shirohana cultivar derived from Chinese lotus) cultivated in Ishikawa Prefecture were investigated in comparison with starch from Chinese lotus cultivated in Ibaraki Prefecture. RVA analysis indicated starch from Kaga lotus had high peak viscosity compared with starch from Ibaraki lotus. Morphological observations showed that starch granules from Kaga lotus had a longer shape than starch granules from the Ibaraki lotus. The amylose content of starch from Kaga lotus was 11.3%, while that from Ibaraki lotus was 14.7%. The phosphorus content was comparable in both starches. Taken together, these results suggest the lower amylose content of starch from Kaga lotus contributes to its high stickiness.