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Bifidobacterium pseudocatenulatum grows well in the early stages of cultivation in medium containing sucrose (Suc), whereas its growth in medium containing the analogue disaccharide N -acetylsucrosamine (SucNAc) tends to exhibit a considerable delay. To elucidate the cause of this phenomenon, we investigated the proliferation pattern of B. pseudocatenulatum in medium containing D-glucose (Glc) and SucNAc and identified the enzyme that degrades this disaccharide. We found that B. pseudocatenulatum initially proliferates by assimilating Glc, with subsequent growth based on SucNAc assimilation depending on production of the β-fructofuranosidase, which can hydrolyze SucNAc, after Glc is completely consumed. Thus, B. pseudocatenulatum exhibited a diauxic growth pattern in medium containing Glc and SucNAc. In contrast, when cultured in medium containing Glc and Suc, B. pseudocatenulatum initially grew by degrading Suc via the phosphorolysis activity of Suc phosphorylase, which did not react to SucNAc. These observations indicate that B. pseudocatenulatum proliferates by assimilating Suc and SucNAc via different pathways. The β-fructofuranosidase of B. pseudocatenulatum exhibited higher hydrolytic activity against several naturally occurring Suc-based tri- or tetrasaccharides than against Suc, suggesting that this enzyme actively catabolizes oligosaccharides other than Suc. [ABSTRACT FROM AUTHOR]

In this study, all four stereoisomers of tryptoline or tetrahydro-β-carboline were synthesized in high yields by the catalyst-free amidation of methyl ester using methylamine under mild conditions. All isomers of the obtained amide and the precursor methyl ester were subjected to cell viability measurements on HeLa cells. The results indicated that the stereochemistry of the derivatives is clearly related to cell viability. [ABSTRACT FROM AUTHOR]

Bacterial strain NYT501, which we previously isolated from soil, was identified as Stenotrophomonas maltophilia, and it was confirmed that this strain produces an intracellular β -N-acetylhexosaminidase exhibiting transglycosylation activity. Several properties of this enzyme were characterized using a partially purified enzyme preparation. Using N, N′-diacetylchitobiose (GlcNAc) and N, N′, N″-triacetylchitotriose (GlcNAc) as substrates and dried cells of this bacterium as a whole-cell catalyst, chitin oligosaccharides of higher degrees of polymerization were synthesized. (GlcNAc) was generated from (GlcNAc) as the major transglycosylation product, and a certain amount of purified sample of the trisaccharide was obtained. By contrast, in the case of the reaction using (GlcNAc) as a substrate, the yield of higher-degree polymerization oligosaccharides was comparatively low. [ABSTRACT FROM AUTHOR]

Chitin oligosaccharide deacetylase (COD) from bacteria that have been examined so far typically comprise two carbohydrate-binding domains (CBDs) and one polysaccharide deacetylase domain. In contrast,Shewanella balticaATCC BAA-1091 COD (Sb-COD) has only one CBD, yet exhibits chitin-binding properties and substrate specificities similar to those of other CODs. [ABSTRACT FROM PUBLISHER]

Using D-glucurono-6,3-lactone (GlcL) as a starting material, sucuronamide (SucAm) was synthesized in two steps using a chemoenzymatic method. In the first chemical synthesis, D-glucuronamide (GlcAm) was synthesized by treating GlcL with NH3 in ethanol. In the second enzymatic synthesis exploiting transfructosylation activity of β-fructofuranosidase, which is inhibited by D-glucose, from Arthrobacter sp. K-1, addition of the sucrose non-assimilating yeast, Saccharomyces bisporus NBRC1131, to the reaction mixture increased SucAm production from GlcAm and sucrose. The overall molar yield of this disaccharide after purification by column chromatography and crystallization was 34.7%. [ABSTRACT FROM AUTHOR]

Enzymatic synthesis of trisaccharides fromN-acetylsucrosamine and lactose utilizing the transgalactosylation activity ofAspergillus oryzaeβ-galactosidase provided two reaction products. Structure analyses by various 2D NMR spectroscopy and MS indicated that the products were β-D-fructofuranosyl β-D-galactopyranosyl-(1→6)-2-acetamido-2-deoxy-α-D-glucopyranoside and β-D-galactopyranosyl-(1→6)-β-D-fructofuranosyl-(2↔1)-2-acetamido-2-deoxy-α-D-glucopyranoside. Moreover,J-resolved-HMBC experiments indicated that the conformations around the glycosidic bonds of these trisaccharides were very similar. Examination about the pH and thermal stabilities of the glycosidic bonds in the GlcNAc–Fru moiety of the two trisaccharides indicated apparent difference. [ABSTRACT FROM AUTHOR]

Two kinds of oligosaccharides, N-acetylraffinosamine (RafNAc) and N-acetylplanteosamine (PlaNAc), were synthesized from N-acetylsucrosamine and melibiose using the transgalactosylation activity of Aspergillus niger α-galactosidase. RafNAc and PlaNAc are novel trisaccharides in which D-glucopyranose residues in raffinose (Raf) and planteose are replaced with N-acetyl-D-glucosamine. These trisaccharides were more stable in acidic solution than Raf. RafNAc was hydrolyzed more rapidly than Raf by α-galactosidase of green coffee bean. In contrast, RafNAc was not hydrolyzed by Saccharomyces cerevisiae invertase, although Raf was hydrolyzed well by this enzyme. These results indicate that the physicochemical properties and steric structure of RafNAc differ considerably from those of Raf. [ABSTRACT FROM AUTHOR]

Two strains [BL21(DE3) and HMS174(DE3)] of Escherichia coli harboring the recombinant chitinase expression plasmid pVP-Chi, which contains Vibrio parahaemolyticus chitinase gene with an attached signal sequence, were prepared. These E. coli transformants produced a large amount of recombinant chitinase, which hydrolyzes chitin to yield di- N-acetylchitobiose (GlcNAc), under the presence of isopropyl-1-thio-β-D-galactopyranoside (IPTG), and secreted the enzyme into their culture fluid with the aid of the signal peptide. Cultivation of these E. coli transformants in Luria-Bertani medium containing squid pen β-chitin and IPTG gave rise to the decomposition of this polysaccharide and the accumulation of (GlcNAc) in the culture fluid. Through these experiments, we confirmed that the use of strain HMS174(DE3) was preferable for the stable accumulation of (GlcNAc) in the culture fluid during cultivation owing to lower (GlcNAc) assimilation compared to BL21(DE3). Next, using E. coli HMS174(DE3) transformants, we conducted saccharification of different forms (fluffy fiber, flake, and powder) of β-chitin samples prepared from squid pens in Bacterion-N-KS(B)K medium containing 2 % of each sample under the presence of IPTG. In these experiments, (GlcNAc) was isolated with a more than 20 % stoichiometric yield from each culture supernatant through charcoal column chromatography followed by recrystallization. [ABSTRACT FROM AUTHOR]

The X-ray crystal structure of chitin oligosaccharide deacetylase from Vibrio parahaemolyticus ( Vp -COD) was determined at an 1.35 Å resolution. The amino acid sequence and structure of Vp -COD show that the enzyme comprises one polysaccharide deacetylase domain (PDD) and two carbohydrate-binding domains (CBDs). On the basis of a chitin-binding assay with Vp -COD and its CBDs-deleted mutant, it was confirmed that CBDs can adhere to chitin. The catalytic activity of the CBDs-deleted mutant was only mildly depressed compared with that of Vp -COD, indicating that CBDs are unlikely to affect the configuration of the active center residues in active site of PDD. [ABSTRACT FROM AUTHOR]

Vibrio parahaemolyticus KN1699 produces 4-O-(N-acetyl-β-d-glucosaminyl)-d-glucosamine (GlcNAc-GlcN) as a major end product from chitin using two extracellular hydrolases: glycoside hydrolase family 18 chitinase, which produces (GlcNAc)2 from chitin, and carbohydrate esterase (CE) family 4 chitin oligosaccharide deacetylase (COD), which hydrolyzes the N-acetyl group at the reducing-end GlcNAc residue of (GlcNAc)2. In this study, we clarified that this heterodisaccharide functions as an inducer of the production of the two above-mentioned chitinolytic enzymes, particularly chitinase. Similar results for chitinase production were obtained with other chitin-decomposing Vibrio strains harboring the CE family 4 COD gene; however, such an increase in chitinase production was not observed in chitinolytic Vibrio strains that did not harbor the COD gene. These results suggest that GlcNAc-GlcN is a unique inducer of chitinase production in Vibrio bacteria that have the COD-producing ability and that the COD involved in the synthesis of this signal compound is one of the key enzymes in the chitin catabolic cascade of these bacteria. [ABSTRACT FROM PUBLISHER]

The article discusses a study which examines the production, secretion and purification of a recombinant Vibrio parahaemolyticus chitinase by Escherichia coli. An open reading frame encoding the chitinase gene and its signal sequence was cloned from V. parahaemolyticus KN1699 genome. An expression plasmid containing the chitinase gene was introduced into Escherichia coli cells, producing recombinant chitinase.

An open reading frame (ORF) encoding chitin oligosaccharide deacetylase ( Pa-COD) gene and its signal sequence was cloned from the Vibrio parahaemolyticus KN1699 genome and its sequence was analyzed. The ORF encoded a 427 amino acid protein, including the 22 amino acid signal sequence. The deduced amino acid sequence was highly similar to several bacterial chitin oligosaccharide deacetylases in carbohydrate esterase family 4. An expression plasmid containing the gene was constructed and inserted into Escherichia coli cells and the recombinant enzyme was secreted into the culture medium with the aid of the signal peptide. The concentration of the recombinant enzyme in the E. coli culture medium was 150 times larger than that of wild-type enzyme produced in the culture medium by V. parahaemolyticus KN1699. The recombinant enzyme was purified to homogeneity from culture supernatant in an overall yield of 16%. Substrate specificities of the wild-type and the recombinant enzymes were comparable. [ABSTRACT FROM AUTHOR]

Abstract: Compared with algal and cyanobacterial cytochrome c 6, cytochrome c 6A from higher plants contains an additional loop of 12 amino acid residues. We have determined the first crystal structure of cytochrome c 6A from Arabidopsis thaliana at 1.5Å resolution in order to help elucidate its function. The overall structure of cytochrome c 6A follows the topology of class I c-type cytochromes in which the heme prosthetic group covalently binds to Cys16 and Cys19, and the iron has octahedral coordination with His20 and Met60 as the axial ligands. Two cysteine residues (Cys67 and Cys73) within the characteristic 12 amino acids loop form a disulfide bond, contributing to the structural stability of cytochrome c 6A. Our model provides a chemical basis for the known low redox potential of cytochrome c 6A which makes it an unsuitable electron carrier between cytochrome b 6 f and PSI. [Copyright &y& Elsevier]

The article presents a study which analyzes the physicochemical characteristic and structure of cytochrome c554 from Vibrio parahaemolyticus strain RIMD2210633. The study compared the terrtiary structure and physicochemical properties of Vibrio cytochrome c554 with Pseudomonas cytochrome c552 by expressing the cytochrome c554 which was expressed in Escherichia coli. The results showed that both cytochromes can denote an electron to an electron carrier.

The article presents a study regarding the physicochemical properties and crystal structure of diatom cytochrome(cyt) c6 coming from Phaeodactylum tricornutum (P. tricornutum). It compares the algal and cyanobacterial cyt c6 P. tricornutum and used 1.5 A resolution. Results of the study reveal that Pct6 structural stability is higher compared to red algal cyt c6. It also notes that a salt bridge is attributed to structural stability.