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Objectives: Catalytic promiscuity, or the ability to catalyze a secondary reaction, provides new opportunities for industrial biocatalysis by expanding the range of biocatalytic reactions. Some nitrilases converting nitriles to amides, referred to as the secondary activity, show great potential for amides production. And our goal was exploiting the amide-forming potential of nitrilases. Results: In this study, we characterized and altered the secondary activity of nitrilase from Acidovorax facilis 72 W (Nit72W) towards different substrates. We increased the secondary activity of Nit72W towards 2-cyanopyridine by 196-fold and created activity toward benzonitrile and p-nitrophenylacetonitrile by modifying the active pocket. Surprisingly, the best mutant, W188M, completely converted 250 mM 2-cyanopyridine to more than 98% 2-picolinamide in 12 h with a specific activity of 90 U/mg and showed potential for industrial applications. Conclusions: Nit72W was modified to increase its secondary activity for the amides production, especially 2-picolinamide. [ABSTRACT FROM AUTHOR]

Objectives: To investigate the lipase-catalyzed synthesis of high purity sn-1/3 and sn-2 monoacylglycerols (1/3-MAG and 2-MAG) of different fatty acids (FAs). Results: The 1/3-MAGs of three FAs (16:0, 17:0, 16:1) were synthesized using lipase-catalyzed esterification of glycerol with FAs. The 2-MAGs were obtained from the ethanolysis of synthetic triacylglycerols using sn-1,3 regiospecific lipase. The effects of lipase types, substrate ratio, temperature, reaction time and lipase load on the MAG conversion were studied. Under the optimal conditions, high purities (96.74%, 95.44%, 92.96%) with acceptable isolated yields (51.00%, 54.28%, 46.00%) were obtained for 1/3-16:0-MAG, 1/3-17:0-MAG, and 1/3-16:1-MAG, respectively. For 2-16:0-MAG, 2-17:0-MAG, and 2-16:1-MAG, the purities were 92.64, 95.04, and 96.48%, with isolated yields of 50.64, 52.16, and 26.12%, respectively. The molecular structures of the synthetic compounds were confirmed by 1H NMR, and MS and the melting points were characterized by DSC. Conclusions: High purity MAG isomers can be synthesized via lipase-catalyzed reactions to be building blocks for production of functional lipids, the melting points of which are largely governed by the hydrophobic interactions among the alkanyl chains. [ABSTRACT FROM AUTHOR]

Two single gene cassettes, each containing one of the individual gene (γ-glutamylcysteine synthetase gene GSH1 or glutathione synthetase gene GSH2), were constructed under the control of alcohol dehydrogenase ( ADH1) promoter and their respective native terminators. The recombinant plasmids constructed with Kan or Hyg as the selective markers and were transformed into Saccharomyces cerevisiae separately and jointly. Three engineered strains, GSH1-enhanced strain S.TS013/GSH1, GSH2-enhanced strain S.TS013/GSH2 and GSH1+GSH2 double-enhanced strain S.TS013/GSH1+GSH2, were constructed. Glutathione production using the recombinant strains to improve was then determined. By the cell dosage proportions of two engineered strains (S.TS013/GSH1, S.TS013/GSH2) and a two-stage reaction, GSH productivity increased by 84 and 59 % over that of the host strain and the S.TS013/GSH1+GSH2 strain, respectively. [ABSTRACT FROM AUTHOR]

Cell surface engineering is a promising strategy for the molecular breeding of whole-cell biocatalysts. By using this strategy, yeasts can be constructed by the cell surface display of functional proteins; these yeasts are referred to as arming yeasts. Because reactions using arming yeasts as whole-cell biocatalysts occur on the cell surface, materials that cannot enter the cell can be used as reaction substrates. Numerous arming yeasts have therefore been constructed for a wide range of uses such as biofuel production, synthesis of valuable chemicals, adsorption or degradation of environmental pollutants, recovery of rare metal ions, and biosensors. Here, we review the science of yeast cell surface modification as well as current applications and future opportunities. [ABSTRACT FROM AUTHOR]

Improvements in current strategies for carrier-based immobilisation have been developed using hetero-functionalised supports that enhance the binding efficacy and stability through multipoint attachment. New commercial resins (Sepabeads) exhibit improved protein binding capacity. Novel methods of enzyme self-immobilisation have been developed (CLEC, CLEA, Spherezyme), as well as carrier materials (Dendrispheres), encapsulation (PEI Microspheres), and entrapment. Apart from retention, recovery and stabilisation, other advantages to enzyme immobilisation have emerged, such as enhanced enzyme activity, modification of substrate selectivity and enantioselectivity, and multi-enzyme reactions. These advances promise to enhance the roles of immobilisation enzymes in industry, while opening the door for novel applications. [ABSTRACT FROM AUTHOR]

α-Chymotrypsin was chemically modified with methoxypoly(ethylene glycol) (PEG) of different molecular weights (700, 2,000, and 5,000 Da) and the amount of polymer attached to the enzyme was varied systematically from 1 to 9 PEG molecules per enzyme molecule. Upon PEG conjugation, enzyme catalytic turnover ( kcat) decreased by 50% and substrate affinity was lowered as evidenced by an increase in the KM from 0.05 to 0.19 mM. These effects were dependent on the amount of PEG bound to the enzyme but were independent of the PEG size. In contrast, stabilization toward thermal inactivation depended on the PEG molecular weight with conjugates with the larger PEGs being more stable. [ABSTRACT FROM AUTHOR]

Sugar cane bagasse hemicellulose, hydrolyzed by dilute H2SO4, supplemented with mineral salts and 0.5% corn steep liquor, was fermented to l(+)-lactic acid using a newly isolated strain of Bacillus sp. In batch fermentations at 50 °C and pH 5, over 5.5% (w/v) l(+)-lactic acid was produced (89% theoretical yield; 0.9 g lactate per g sugar) with an optical purity of 99.5%. [ABSTRACT FROM AUTHOR]

An NADPH-dependent carbonyl reductase that shows (3R)-selective reducing activity for α-substituted-β-ketoesters was purified from Kluyveromyces marxianus and racemic alkyl 2-substituted-3-oxobutanoates were reduced to the corresponding (2S,3R)- or (2R,3R)-2-substituted-3-hydroxybutanoates with enantiomeric purity (> 99%) and diastereoselectivity (24 ∼ 98%). [ABSTRACT FROM AUTHOR]

A method for determining the reactant concentration and biocatalyst activity during a biocatalysed reaction is described. The analyser functions by monitoring the temperature change of a sample of reaction mixture which is held under adiabatic conditions. Details are given of the successful application of the analyser to the monitoring and control of a fed-batch bioconversion by Rhodococcus ruber NCIMB 40757 of acrylonitrile to 42% (w/w) (4.37 M) ammonium acrylate and the wider application of the analyser to other bioconversions is considered. [ABSTRACT FROM AUTHOR]

An automated CSTR system has been developed to study the stability of immobilised whole cell biocatalysts for ammonium acrylate production. Two potentially useful amidase-active rhodococci have been identified: One possesses an amidase of much higher activity than the other but of much lower stability. The activity and stability of each biocatalyst over a range of temperatures has been determined and used to compare the productivity of the biocatalysts in a bioconversion. [ABSTRACT FROM AUTHOR]

Lipase and newly synthesized gemini-type detergent were mixed in an aqueous phosphate buffer solution and lyophilized. Due to complex formation between the substances it was possible to dissolve or at least finely disperse these preparations in organic media. The lipase-detergent complexes were used to catalyse the irreversible transesterfication of 6-methyl-5-hepten-2-ol (sulcatol) or 2,2-dimethyl-1,3-dioxolane-4-methanol (solketal) with vinyl or isopropenyl carboxylate in organic solvents. The complexes were considerably more active than enzyme powder or the complexes prepared with conventional synthesized detergents in organic media. © Rapid Science Ltd. 1998 [ABSTRACT FROM AUTHOR]