Although many methods for enzyme immobilization have been described in patents and publications, relatively few processes employing immobilized enzymes have been successfully commercialized. The cost of most industrial enzymes is often only a minor component in overall process economics, and in these instances, the additional costs associated with enzyme immobilization are often not justified. More commonly the benefit realized from enzyme immobilization relates to the process advantages that an immobilized catalyst offers, for example, enabling continuous production, improved stability and the absence of the biocatalyst in the product stream. The development and attributes of several established and emerging industrial applications for immobilized enzymes, including high-fructose corn syrup production, pectin hydrolysis, debittering of fruit juices, interesterification of food fats and oils, biodiesel production, and carbon dioxide capture are reviewed herein, highlighting factors that define the advantages of enzyme immobilization.

Phage peptide display libraries are commonly used to select peptides that bind to inorganic surfaces (metals, metal oxides, and semiconductors). These binding peptides can serve as templates to control the nucleation and growth of inorganic nanoparticles in vitro. In this report, we describe the identification of a unique set of sequences that bind to silver and cobalt nanoparticles from a phage peptide display library using a polymerase chain reaction (PCR)‐driven method. The amino acid sequences obtained by the PCR method are a distinct set of sequences that would otherwise be missed using the regular panning method. Peptides identified by the method described here are also shown to function as templates for the synthesis of silver and cobalt platinum nanoparticles.

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Summary Emissions of biogenic volatile organic compounds (bVOCs), are an important element in the global carbon cycle, accounting for a significant proportion of fixed carbon. They contribute directly and indirectly to global warming and climate change and have a major effect on atmospheric chemistry. Plants emit isoprene to the atmosphere in similar quantities to emissions of methane from all sources and each accounts for approximately one third of total VOCs. Although methanotrophs, capable of growth on methane, have been intensively studied, we know little of isoprene biodegradation. Here, we report the isolation of two isoprene‐degrading strains from the terrestrial environment and describe the design and testing of polymerase chain reaction (PCR) primers targeting isoA, the gene encoding the active‐site component of the conserved isoprene monooxygenase, which are capable of retrieving isoA sequences from isoprene‐enriched environmental samples. Stable isotope probing experiments, using biosynthesized 13C‐labelled isoprene, identified the active isoprene‐degrading bacteria in soil. This study identifies novel isoprene‐degrading strains using both culture‐dependent and, for the first time, culture‐independent methods and provides the tools and foundations for continued investigation of the biogeography and molecular ecology of isoprene‐degrading bacteria.

Polysialylated neural cell adhesion molecule (NCAM)is thought to play a critical role in neural development. Polysialylation of NCAM was shown to be achieved by two ␣2,8-polysialyltransferases, ST8Sia IV (PST) and ST8Sia II (STX), which are moderately related to another ␣2,8-sialyltransferase, ST8Sia III. Here we describe that all three ␣2,8-sialyltransferases can utilize oligosaccharides as acceptors but differ in the efficiency of adding polysialic acid on NCAM. First, we found that ST8Sia III can form polysialic acid on the enzyme itself (autopolysialylation) but not on NCAM. These discoveries prompted us to determine if ST8Sia IV and ST8Sia II share the property of ST8Sia III in utilizing low molecular weight oligosaccharides as acceptors. By using a newly established method, we found that ST8Sia IV, ST8Sia II, and ST8Sia III all add oligosialic and polysialic acid on various sialylated N-acetyllactosaminyl oligosaccharides, including NCAM N-glycans, fetuin N-glycans, synthetic sialylated N-acetyllactosamines, and on ␣ 2 -HS-glycoprotein. Our results also showed that monosialyl and disialyl N-acetyllactosamines can serve equally as an acceptor, suggesting that no initial addition of ␣2,8-sialic acid is necessary for the action of polysialyltransferases. Polysialylation of NCAM by ST8Sia IV and ST8Sia II is much more efficient than polysialylation of N-glycans isolated from NCAM. Moreover, ST8Sia IV and ST8Sia II catalyze polysialylation of NCAM much more efficiently than ST8Sia III. These results suggest that no specific acceptor recognition is involved in polysialylation of low molecular weight sialylated oligosaccharides, whereas the enzymes exhibit pronounced acceptor specificities if glycoproteins are used as acceptors.

Human corneal N-acetylglucosamine 6-O-sulfotransferase (hCGn6ST) has been identified by the positional candidate approach as the gene responsible for macular corneal dystrophy (MCD). Because of its high homology to carbohydrate sulfotransferases and the presence of mutations of this gene in MCD patients who lack sulfated keratan sulfate in the cornea and serum, hCGn6ST protein is thought to be a sulfotransferase that catalyzes sulfation of GlcNAc in keratan sulfate. In this report, we analyzed the enzymatic activity of hCGn6ST by expressing it in cultured cells. A lysate prepared from HeLa cells transfected with an intact form of hCGn6ST cDNA or culture medium from cells transfected with a secreted form of hCGn6ST cDNA showed an activity of transferring sulfate to C-6 of GlcNAc of synthetic oligosaccharide substrates in vitro. When hCGn6ST was expressed together with human keratan sulfate Gal-6-sulfotransferase (hKSG6ST), HeLa cells produced highly sulfated carbohydrate detected by an anti-keratan sulfate antibody 5D4. These results indicate that hCGn6ST transfers sulfate to C-6 of GlcNAc in keratan sulfate. Amino acid substitutions in hCGn6ST identical to changes resulting from missense mutations found in MCD patients abolished enzymatic activity. Moreover, mouse intestinal GlcNAc 6-O-sulfotransferase had the same activity as hCGn6ST. This observation suggests that mouse intestinal GlcNAc 6-O-sulfotransferase is the orthologue of hCGn6ST and functions as a sulfotransferase to produce keratan sulfate in the cornea.

Poly-N-acetyllactosamine is a unique carbohydrate composed of N-acetyllactosamine repeats and provides the backbone structure for additional modifications such as sialyl Le x . Poly-N-acetyllactosamines in mucintype O-glycans can be formed in core 2 branched oligosaccharides, which are synthesized by core 2 ␤-1,6-Nacetylglucosaminyltransferase.Using a ␤-1,4-galactosyltransferase (␤4Gal-TI) present in milk and the recently cloned ␤-1,3-N-acetylglucosaminyltransferase, the formation of poly-N-acetyllactosamine was found to be extremely inefficient starting from a core 2 branched oligosaccharide, GlcNAc␤136-(Gal␤133)GalNAc␣3 R. Since the majority of synthesized oligosaccharides contained N-acetylglucosamine at the nonreducing ends, galactosylation was judged to be inefficient, prompting us to test novel members of the ␤4Gal-T gene family for this synthesis. Using various synthetic acceptors and recombinant ␤4Gal-Ts, ␤4Gal-TIV was found to be most efficient in the addition ofasinglegalactoseresiduetoGlcNAc␤136(Gal␤133)GalNAc␣3 R. Moreover, ␤4Gal-TIV, together with ␤-1,3-Nacetylglucosaminyltransferase, was capable of synthesizing poly-N-acetyllactosamine in core 2 branched oligosaccharides. On the other hand, ␤4Gal-TI was found to be most efficient for poly-N-acetyllactosamine synthesis in N-glycans. In contrast to ␤4Gal-TI, the efficiency of ␤4Gal-TIV decreased dramatically as the acceptors contained more N-acetyllactosamine repeats, consistent with the fact that core 2 branched O-glycans contain fewer and shorter poly-N-acetyllactosamines than N-glycans in many cells. These results, as a whole, indicate that ␤4Gal-TIV is responsible for poly-Nacetyllactosamine synthesis in core 2 branched O-glycans.Mucin-type O-glycans are present in a wide variety of cells and play various roles in different cells. Mucin-type glycoproteins are also present in the plasma membrane, and they are often involved in cell-cell interaction (1). For example, O-glycans present in eggs were shown to be a receptor for both mouse and sea urchin (2, 3). In granulocytes, monocytes, and certain T lymphocytes, mucin-type O-glycans can carry sialyl Le x , NeuNAc␣233Gal␤134(Fuc␣133)GlcNAc3 R, at their termini (4 -6). Sialyl Le x and its sulfated form are ligands for E-, P-, and L-selectin (7-11). Importantly, these selectins, in particular P-and L-selectin, preferentially bind to sialyl Le x in a limited number of mucin-type glycoproteins such as PSGL-1 (for P-selectin) and GlyCAM-1 and CD34 (for L-selectin) (12-14). As shown previously, sialyl Le x and its derivatives of O-glycans in blood cells can be only formed on core 2 branches, Gal␤134GlcNAc␤136(Gal␤133)GalNAc␣3 R (4, 5). Recent studies demonstrate that sialyl Le x and sialyl Le a in core 2 branches are highly correlated to tumor invasion and vessel invasion of colon carcinomas (15), probably because tumor cells utilize selectin-carbohydrate interaction for their adhesion.In patients with immunodeficiency such as Wiskott-Aldrich syndrome, AIDS, and leukemia, leukocytes in the peripheral blood...

Treatment of the peracetylated ethyl dithioacetals of D-glucose, D-galactose, and D-mannose with acetyl chloride and boron trifluoride diethyl etherate at reflux yields the known acyclic 1-chloro-1-(ethylthio) derivatives. These compounds are shown to effectively glycosylate a variety of carbohydrate acceptors using silver trifluoromethanesulfonate as the promotor, affording stereospecifically the corresponding acyclic O,S-acetals in good to excellent yield. Furthermore, following deacetylation, treatment of these O,S-acetals with a mixture of mercuric salts in either methanol or dimethylformamide gives rise to disaccharides terminating in D-furanosyl residues.