Biocatalysis Research Papers - Academia.edu (original) (raw)

High-level composite, ab initio and density functional theory (DFT) procedures have been employed to study O–H bond dissociation energies (BDEs), as well as radical stabilization energies (RSEs) in the oxygen-centred radicals that are... more

High-level composite, ab initio and density functional theory (DFT) procedures have been employed to study O–H bond dissociation energies (BDEs), as well as radical stabilization energies (RSEs) in the oxygen-centred radicals that are formed in the dissociation of the O–H bonds. Benchmark values are provided by Wn results up to W3.2 and W4.x. We are able to recommend revised BDE values for FO–H (415.6 ± 3 kJ mol–1), MeC(O)O–H (459.8 ± 6 kJ mol–1) and CF3CH2O–H (461.9 ± 6 kJ mol–1) on the basis of high-level calculations. We find that Gn-type procedures are generally reliable and cost-effective, and that some contemporary functionals and double-hybrid DFT procedures also provide adequate O–H BDEs/RSEs. We note that the variations in the O–H BDEs are associated with variations in the stabilities of not only the radicals but also the closed-shell precursor molecules. Most substituents destabilize both species, with σ-electron-withdrawing groups having larger destabilizing effects, whil...

This paper examined the effect of several pyridinium and imidazolium-based ionic liquids (ILs) on the protease stability in aqueous solutions. In general, the enzyme was found quite active at low concentrations of hydrophilic ILs. In... more

This paper examined the effect of several pyridinium and imidazolium-based ionic liquids (ILs) on the protease stability in aqueous solutions. In general, the enzyme was found quite active at low concentrations of hydrophilic ILs. In aqueous environment, the enzyme was stabilized by the kosmotropic anions (such as CF3COO− and CH3COO−) and chaotropic cations (such as [BuPy]+ and [EMIM]+), but was destabilized by chaotropic anions (such as tosylate and BF4−) and kosmotropic cations (such as [BMIM]+).Hydrophilic ionic liquids dissociate into individual ions in water. The effect of individual ions on the enzyme activity follows the Hofmeister series: kosmotropic anions and chaotropic cations stabilize the enzyme.

Immobilization of β-GAL could represent an important driving force for the development of lactose hydrolysis and galacto-oligosaccharides (GOS) synthesis technology. The enzymatic hydrolysis of lactose is one of the most important... more

Immobilization of β-GAL could represent an important driving force for the development of lactose hydrolysis and galacto-oligosaccharides (GOS) synthesis technology. The enzymatic hydrolysis of lactose is one of the most important biotechnological processes in the food industry. It is carried out by β-d-galactosidase (also named lactase, EC 3.2.1.108), an enzyme used in several applications for hydrolysis of lactose from milk or whey. The main benefit of lactose-hydrolyzed products is that they overcome lactose intolerance, present in more than half of population of the world. Manufacture of lactose-free milk and dairy products is important to allow consumption of such foods for people (especially children) with intestinal lactase deficiency, resulting in lactose intolerance (Panesar et al., 2006). Another important result of the lactose hydrolysis process is the increased sweetening power and solubility of the obtained monosaccharides, which can generate new applications,

Enzymes, being remarkable catalysts, are capable of accepting a wide range of complex molecules as substrates and catalyze a variety of reactions with a high degree of chemo-, stereo- and regioselectivity in most of the reactions.... more

Enzymes, being remarkable catalysts, are capable of accepting a wide range of complex molecules as substrates and catalyze a variety of reactions with a high degree of chemo-, stereo- and regioselectivity in most of the reactions. Biocatalysis can be used in both simple and complex chemical transformations without the need for tedious protection and deprotection chemistry that is very common in traditional organic synthesis. This current review highlights the applicability of one class of biocatalysts viz.…

The advances in studies of trace metal speciation and bioavailability since Mark Florence’s 1982 review of the topic, published in Talanta, have been comprehensively reviewed. While the relative merits of kinetic and equilibrium... more

The advances in studies of trace metal speciation and bioavailability since Mark Florence’s 1982 review of the topic, published in Talanta, have been comprehensively reviewed. While the relative merits of kinetic and equilibrium approaches are still being determined, advances in the applications of stripping voltammetry, including the application of microelectrodes and an appreciation of detection windows in both CSV and ASV, have been matched by the introduction of new dynamic techniques including diffusive gradients in thin films (DGTs), permeation liquid membranes (PLMs), and improved applications of chelating resins. There have also been improvements in equilibrium techniques such as ion-selective electrodes and Donnan dialysis. The ability of geochemical speciation models to predict metal complexation by natural organic matter has greatly improved, yet the models still require validation against field measurements. More reliable and relevant bioassays have been developed using ...

The ability of peroxidases and laccases enzymes to treat organic pollutants is reviewed. Enzymatic methods generally have low energy requirements, are easy to control, can operate over a wide range of conditions and have a minimal... more

The ability of peroxidases and laccases enzymes to treat organic pollutants is reviewed. Enzymatic methods generally have low energy requirements, are easy to control, can operate over a wide range of conditions and have a minimal environmental impact. Peroxidases and laccases have broad substrate specificities and can catalyze the oxidation of a wide range of toxic organic compounds. The results show that an enzymatic oxidation can diminish the toxicity of some polycyclic aromatic hydrocarbons (PAHs), phenols, organophosphorus pesticides and azo dyes in laboratory and some field conditions. Due to the hydrophobicity and low aqueous solubility of these substrates, reactions are usually performed in the presence of organic solvents. However, it was detected that organic solvents can provoke enzyme denaturation, unfavorable substrate partition, inhibition or stabilization of enzyme–substrate complexes, depending on the enzyme, substrate and organic solvent used. Strategies to overcome these problems are proposed. Additionally, the low stability of heme-containing peroxidases to hydrogen peroxide, the low reaction rates of laccases, the mediators toxicity, the limited availability and high costs of these enzymes are other limitations detected for commercial applications. Due to field reaction conditions are more complex than laboratory conditions efforts have to be made to achieve the cheap overproduction of these biocatalysts in heterologous hosts and also their modification by chemical means or protein engineering to obtain more robust and active enzymes.

ABSTRACT Enzyme catalysis: Three dehydrogenases have been engineered to self-assemble into a hydrogel that supports a synthetic metabolic network. The new catalytic biomaterial was used as an anode modification in two enzymatic... more

ABSTRACT Enzyme catalysis: Three dehydrogenases have been engineered to self-assemble into a hydrogel that supports a synthetic metabolic network. The new catalytic biomaterial was used as an anode modification in two enzymatic biobatteries capable of the complete oxidation of methanol to CO(2) .

Development of artificial enzymes designed for industrial plants that could convert carbon dioxide into carbonates, with the ultimate aim of reducing CO2 emissions. Enzymes are biological catalysts that accelerate chemical reactions, such... more

Development of artificial enzymes designed for industrial plants that could convert carbon dioxide into carbonates, with the ultimate aim of reducing CO2 emissions. Enzymes are biological catalysts that accelerate chemical reactions, such as the conversion of gaseous carbon dioxide (CO2) into carbonates. Carbonates are the basic component of coral reefs, mollusc shells, geological platforms and kidney stones. Although naturally occurring enzymes would be ideal for converting human-generated CO2 emissions into carbonates, they are generally incapable of coping with the extreme conditions of industrial plants. Ernesto and colleagues are now developing artificial enzymes that can withstand the harsh environments of industrial plants while accelerating chemical reactions. His team ultimately aims to create a clean, cheap, practical and socially responsible solution for global warming by reducing CO2 emissions. "We believe that our novel artificial enzymes will be the first tailor-made enzymes for industrial plants to produce carbonates," says Dr Hernandez. So far, Dr Hernandez and his colleagues have built an artificial environment composed of chimney-like equipment, measuring 1.5 metres in height and 15 centimetres in diameter, that mimics the smoke released by power plants. Using the artificial environment, the researchers will ensure that their artificial enzymes can function properly under extreme conditions consisting of hot, corrosive, poisonous and sticky smoke as well as soot and other gases produced by power plants. The team is basing the development of its artificial enzyme on naturally occurring carbonic anhydrase (CA), which accelerates the conversion of CO2 into carbonates. Carbonic anhydrase is capable of turning CO2 molecules into carbonates at a rate of one million molecules per second. However, "the enzyme's CO2 conversion rate slows down dramatically under industrial conditions," Dr Hernandez points out. He and his colleagues are now engineering artificial enzymes based on natural CA, using directed evolution techniques. Their first step involves the creation of a library of diverse genes that encode for carbonic anhydrases. "This library includes sequences of unique forms of carbonic anhydrases recently found near deep-ocean chimneys (hydrothermal vents)," says Dr Hernandez. The team plans to modify and multiply the genes encoding for carbonic anhydrases using a molecular technique called random mutagenesis. The researchers will then place the mutated genes in the artificial environment to see which ones are most effective at converting carbon dioxide into carbonates. The best mutations will then be put through the modification and multiplication processes again. The researchers will repeat the whole process until they have isolated a mutated gene encoding for recombinant carbonic anhydrase that can convert CO2 into carbonates under industrial conditions. With the help of artificial enzymes, CO2-converted carbonates could be used in everything from baking soda and chalk to Portland cement and lime manufacturing.

The study characterized heterogeneous biocatalyst synthesized from sucrose, saw dust, and chicken egg shells using Fourier Transform Infrared (FTIR) spectroscopy coupled with Attenuated Total Reflectance (ATR) technique. Acidic sulphonate... more

The study characterized heterogeneous biocatalyst synthesized from sucrose, saw dust, and chicken egg shells using Fourier Transform Infrared (FTIR) spectroscopy coupled with Attenuated Total Reflectance (ATR) technique. Acidic sulphonate (–SO3H) groups were more visible in the spectrum generated for carbonized and sulphonated sucrose than in carbonized and sulphonated saw dust. This was highlighted further by the significantly higher conversion percentage achieved for sulphonated sucrose (62.5%) than sulphonated saw dust (46.6%) during esterification of expired sunflower oil (p=0.05). The spectra for calcinated egg shells also showed that the most active form of calcium oxide was produced at calcination temperature of 1000°C. This was confirmed in the single-step transesterification reaction in which calcium oxide generated at 1000°C yielded the highest biodiesel (87.8%) from expired sunflower oil. The study further demonstrated the versatility of the FTIR technique in qualitative ...

The author discusses the benefits for forensic science from greater engagement with basic and other applied areas of chemistry and gives examples of how his organization, the Australian Federal Police, have partnered with academia and... more

The author discusses the benefits for forensic science from greater engagement with basic and other applied areas of chemistry and gives examples of how his organization, the Australian Federal Police, have partnered with academia and others to promote the use of chemistry in areas of trace evidence, illicit drugs, fingerprint detection, and explosives.

Tyrosine hydroxylase, an iron containing tetrahydrobiopterin dependent monooxygenase (tyrosine 3-monooxygenase; EC 1.14.16.2), catalyzes the rate-limiting step in which l-dopa is formed from the substrate l-tyrosine. l-Dopa concentration... more

Tyrosine hydroxylase, an iron containing tetrahydrobiopterin dependent monooxygenase (tyrosine 3-monooxygenase; EC 1.14.16.2), catalyzes the rate-limiting step in which l-dopa is formed from the substrate l-tyrosine. l-Dopa concentration and activity of l-tyrosine hydroxylase enzyme were measured in roots, stem, leaves, pods, and immature seeds of Mucuna pruriens. Immature seeds contained maximum l-dopa content and mature leaves possessed maximum catalytic activity of tyrosine hydroxylase. Tyrosine hydroxylase from leaf homogenate was characterized as a 55 kDa protein by SDS-PAGE and Western-blot analysis with monoclonal mouse IgG2a tyrosine hydroxylase antibody. The conditions for maximum tyrosine hydroxylase activity from the leaf extract were optimized with respect to temperature, pH, cofactor 6-MPH4, and divalent metal ions. The tyrosine hydroxylase from leaf extract possessed a K m value of 808.63 μM for l-tyrosine at 37°C and pH 6.0. The activity of the enzyme was slightly inhibited at 2,000 μM l-tyrosine. Higher concentrations of the cofactor 6-MPH4, however, completely inhibited the synthesis of l-dopa. Tyrosine hydroxylase converted specific monophenols such as l-tyrosine (808.63 μM) and tyramine (K m 1.1 mM) to diphenols l-dopa and dopamine, respectively. Fe(II) activated the enzyme while higher concentration of other divalent metals reduced its activity. For the first time, tyrosine hydroxylase from M. pruriens is being reported in this study.

Enzymes utilize substrate binding energy both to promote ground state association and to selectively lower the energy of the reaction transition state.i The monomeric homing endonuclease I-AniI cleaves with high sequence specificity in... more

Enzymes utilize substrate binding energy both to promote ground state association and to selectively lower the energy of the reaction transition state.i The monomeric homing endonuclease I-AniI cleaves with high sequence specificity in the center of a 20 base-pair DNA target site, with the N-terminal domain of the enzyme making extensive binding interactions with the left (−) side of the target site and the similarly structured C-terminal domain interacting with the right (+) side.ii Despite the approximate two-fold symmetry of the enzyme-DNA complex, we find that there is almost complete segregation of interactions responsible for substrate binding to the (−) side of the interface and interactions responsible for transition state stabilization to the (+) side. While single base-pair substitutions throughout the entire DNA target site reduce catalytic efficiency, mutations in the (−) DNA half-site almost exclusively increase KD and KM*, and those in the (+) half-site primarily decrease kcat*. The reduction of activity produced by mutations on the (−) side, but not mutations on the (+) side, can be suppressed by tethering the substrate to the endonuclease displayed on the surface of yeast. This dramatic asymmetry in the utilization of enzyme-substrate binding energy for catalysis has direct relevance to the redesign of endonucleases to cleave genomic target sites for gene therapy and other applications. Computationally redesigned enzymes that achieve new specificities on the (−) side do so by modulating KM*, while redesigns with altered specificities on the (+) side modulate kcat*. Our results illustrate how classical enzymology and modern protein design can each inform the other.

Enzyme immobilization often achieves reusable biocatalysts with improved operational stability and solvent resistance. However, these modifications are generally associated with a decrease in activity or detrimental modifications in... more

Enzyme immobilization often achieves reusable biocatalysts with improved operational stability and solvent resistance. However, these modifications are generally associated with a decrease in activity or detrimental modifications in catalytic properties. On the other hand, protein engineering aims to generate enzymes with increased performance at specific conditions by means of genetic manipulation, directed evolution and rational design. However, the achieved biocatalysts are generally generated as soluble enzymes, −thus not reusable-and their performance under real operational conditions is uncertain. Combined protein engineering and enzyme immobilization approaches have been employed as parallel or consecutive strategies for improving an enzyme of interest. Recent reports show efforts on simultaneously improving both enzymatic and immobilization components through genetic modification of enzymes and optimizing binding chemistry for site-specific and oriented immobilization. Nonetheless, enzyme engineering and immobilization are usually performed as separate workflows to achieve improved biocatalysts. In this review, we summarize and discuss recent research aiming to integrate enzyme immobilization and protein engineering and propose strategies to further converge protein engineering and enzyme immobilization efforts into a novel "immobilized biocatalyst engineering" research field. We believe that through the integration of both enzyme engineering and enzyme immobilization strategies, novel biocatalysts can be obtained, not only as the sum of independently improved intrinsic and operational properties of enzymes, but ultimately tailored specifically for increased performance as immobilized biocatalysts, potentially paving the way for a qualitative jump in the development of efficient, stable biocatalysts with greater real-world potential in challenging bioprocess applications.

Industrial Biorefineries and White Biotechnology provides a comprehensive look at the increasing focus on developing the processes and technologies needed for the conversion of biomass to liquid and gaseous fuels and chemicals, in... more