Recommendations for performing measurements of apparent equilibrium constants of enzyme-catalyzed reactions and for reporting the results of these measurements.

The measurement of values of apparent equilibrium constants ' for enzyme-catalyzed reactions involve a substantial number of critical details, neglect of which could lead to systematic errors. Here, interferences, impurities in the substances used, and failure to achieve equilibrium are matters of substantial consequence. Careful reporting of results is of great importance if the results are to have archival value.

Kinetics of enzyme-catalysed desymmetrisation of prochiral substrates: product enantiomeric excess is not always constant.

The kinetics of enzymatic desymmetrisation were analysed for the most common kinetic mechanisms: ternary complex ordered (prochiral ketone reduction); ping-pong second (ketone amination, diol esterification, desymmetrisation in the second half reaction); ping-pong first (diol ester hydrolysis) and ping-pong both (prochiral diacids). For plausible values of enzyme kinetic parameters, the product enantiomeric excess (ee) can decline substantially as the reaction proceeds to high conversion. For example, an ee of 0.

Thermodynamic Favorability of End Products of Anaerobic Glucose Metabolism.

The eQuilibrator component contribution method allows calculation of the overall Gibbs energy changes for conversion of glucose to a wide range of final products in the absence of other oxidants. Values are presented for all possible combinations of products with up to three carbons and selected others. The most negative Gibbs energy change is for the formation of graphite and water (-499 kJ mol) followed by CH and CO (-430 kJ mol), the observed final products of anaerobic digestion.

Highly Active Protein Surfaces Enabled by Plant-Based Polyphenol Coatings.

Proteins represent complex biomolecules capable of wide-ranging but also highly specific functionalities. Their immobilization on material supports can enable broad applications from sensing and industrial biocatalysis to biomedical interfaces and materials. We demonstrate the advantages of using aqueous-processed cross-linked polyphenol coatings for immobilizing proteins, including IgG, avidin, and various single and multidomain enzymes on diverse materials, to enable active biofunctional structures (e.

An empirical analysis of enzyme function reporting for experimental reproducibility: Missing/incomplete information in published papers.

A key component of enzyme function experiments is reporting of considerable metadata, to allow other researchers to replicate, interpret properly or use fully the results. This paper evaluates the completeness of enzyme function data reporting for reproducibility. We present a detailed examination of 11 recent papers (and their supplementary material) from two leading journals.

Two-dimensional H and H-detected NMR study of a heterogeneous biocatalyst using fast MAS at high magnetic fields.

Nuclear magnetic resonance (NMR) is a powerful tool for investigating atomic-scale structure in heterogeneous or composite materials where long-range order is absent. In this work solid-state H and H-detected NMR experiments were performed with fast magic angle spinning (ν = 75 kHz) and at high magnetic fields (B = 20 T) and used to gain structural insight into a heterogeneous biocatalyst consisting of an enzyme, human carbonic anhydrase II (hCA II), covalently immobilized on epoxy-functionalized silica. Two-dimensional H-H NOESY-type correlation experiments were able to provide information on H environments in silica, epoxy-silica and the immobilized enzyme.

STRENDA DB: enabling the validation and sharing of enzyme kinetics data.

Standards for reporting enzymology data (STRENDA) DB is a validation and storage system for enzyme function data that incorporates the STRENDA Guidelines. It provides authors who are preparing a manuscript with a user-friendly, web-based service that checks automatically enzymology data sets entered in the submission form that they are complete and valid before they are submitted as part of a publication to a journal.

Enzymatic Synthesis of Glucose-Based Fatty Acid Esters in Bisolvent Systems Containing Ionic Liquids or Deep Eutectic Solvents.

Sugar fatty acid esters (SFAEs) are biocompatible nonionic surfactants with broad applications in food, cosmetic, and pharmaceutical industries. They can be synthesized enzymatically with many advantages over their chemical synthesis. In this study, SFAE synthesis was investigated by using two reactions: (1) transesterification of glucose with fatty acid vinyl esters and (2) esterification of methyl glucoside with fatty acids, catalyzed by Lipozyme TLIM and Novozym 435 respectively.

Fluid-dynamical model for antisurfactants.

We construct a fluid-dynamical model for the flow of a solution with a free surface at which surface tension acts. This model can describe both classical surfactants, which decrease the surface tension of the solution relative to that of the pure solvent, and antisurfactants (such as many salts when added to water, and small amounts of water when added to alcohol) which increase it. We demonstrate the utility of the model by considering the linear stability of an infinitely deep layer of initially quiescent fluid.

Ionization basis for activation of enzymes soluble in ionic liquids.

Background: The complex interactions between electrolytes and proteins have been studied for more than a century. However, understanding is not yet complete and does not provide a basis for predicting the activity of enzymes in ionic media. The use of ionic liquids (ILs) as reaction medium has opened up new opportunities for better understanding of the mechanism of enzymatic catalysis.

Supramolecular Fibers in Gels Can Be at Thermodynamic Equilibrium: A Simple Packing Model Reveals Preferential Fibril Formation versus Crystallization.

Low molecular weight gelators are able to form nanostructures, typically fibers, which entangle to form gel-phase materials. These materials have wide-ranging applications in biomedicine and nanotechnology. While it is known that supramolecular gels often represent metastable structures due to the restricted molecular dynamics in the gel state, the thermodynamic nature of the nanofibrous structure is not well understood.

Effect of pore size on the performance of immobilised enzymes.

Porous materials are widely employed as supports in the immobilisation of enzymes. Traditionally macroporous materials with pore diameters >50 nm were believed to be the most suitable support material, ensuring no spatial restrictions upon enzyme molecules entering such large pores. In recent years however, there has been growing emphasis in the use of mesoporous supports with pore diameters ranging between 2 and 50 nm.

Detailed study of precipitation of a poorly water soluble test compound using methodologies as in activity and solubility screening - mixing and automation effects.

Storage of pharmaceutical discovery compounds dissolved in dimethylsulfoxide (DMSO) is commonplace within industry. Often, the DMSO stock solution is added to an aqueous system (e.g.

Isolation breeds naivety: island living robs Australian varanid lizards of toad-toxin immunity via four-base-pair mutation.

Since their introduction to the toad-free Australian continent cane toads (Bufo marinus) have caused a dramatic increase in naïve varanid mortality when these large lizards attempt to feed on this toxic amphibian. In contrast Asian-African varanids, which have coevolved with toads, are resistant to toad toxin. Toad toxins, such as Bufalin target the H1-H2 domain of the α(1) subunit of the sodium-potassium-ATPase enzyme.

Activation of alpha chymotrypsin by three phase partitioning is accompanied by aggregation.

Precipitation of alpha chymotrypsin in the simultaneous presence of ammonium sulphate and t-butanol (three phase partitioning) resulted in preparations which showed self aggregation of the enzyme molecules. Precipitation with increasing amounts of ammonium sulphate led to increasing size of aggregates. While light scattering estimated the hydrodynamic diameter of these aggregates in the range of 242-1124 nm; Nanoparticle tracking analysis (NTA) gave the value as 130-462 nm.

Salt-induced control of supramolecular order in biocatalytic hydrogelation.

Biocatalytic action and specific ion effects are both known to have dramatic effects on molecular self-assembly and hydrogelation. In this paper, we demonstrate that these effects are highly cooperative. Biocatalytic hydrogelation of Fmoc peptides in the presence of salts combines kinetic (through enzymatic catalysis) and thermodynamic (specific ion and protein templating) contributions when applied in combination.

Dramatic specific-ion effect in supramolecular hydrogels.

We report on a pronounced specific-ion effect on the intermolecular and chiral organization, supramolecular structure formation, and resulting materials properties for a series of low molecular weight peptide-based hydrogelators, observed in the presence of simple inorganic salts. This effect was demonstrated using aromatic short peptide amphiphiles, based on fluorenylmethoxycarbonyl (Fmoc). Gel-phase materials were formed due to molecular self-assembly, driven by a combination of hydrogen bonding and π-stacking interactions.

Interaction of counterions with subtilisin in acetonitrile: insights from molecular dynamics simulations.

A recent X-ray structure has enabled the location of chloride and cesium ions on the surface of subtilisin Carlsberg in acetonitrile soaked crystals. (1) To complement the previous study and analyze the system in solution, molecular dynamics (MD) simulations, in acetonitrile, were performed using this structure. Additionally, Cl(-) and Cs(+) ions were docked on the protein surface and this system was also simulated.

BESSICC, a COSMO-RS based tool for in silico solvent screening of biocatalyzed reactions.

Many enzymatic reactions are near-equilibrium reactions. This often limits final yield and hence application of biocatalyzed processes in the industrial production. The most widely applied strategy to overcome this issue is solvent selection.

Examining structure-activity correlations of some high activity enzyme preparations for low water media.

A first study of the comparison of structures of enzymes (by FT-IR and CD) in different high activity (in low water media) preparations is reported. Using chymotrypsin and subtilisin as models, we have studied various factors that distinguish enzyme precipitated and rinsed with propanol (EPRP), crosslinked enzyme aggregates (CLEA), protein coated microcrystals (PCMC) and crosslinked protein coated microcrystals (CLPCMC). The suspensions in organic media were assayed for catalytic activity, and structures were probed by FT-IR and CD measurements.

Enzymatic acylation of starch.

Starch a cheap, abundant and renewable natural material has been chemically modified for many years. The popular modification acylation has been used to adjust rheological properties as well as deliver polymers with internal plasticizers and other potential uses. However the harsh reaction conditions required to produce these esters may limit their use, especially in sensitive applications (foods, pharmaceuticals, etc.

Experimental test of absorption flattening correction for circular dichroism of particle suspensions.

There are well established theoretical models for correction for absorption flattening of circular dichroism (CD) measurements on particle suspensions. However, these have not been directly tested experimentally. We describe a test system with the chiral tris(ethylenediamine)Co(III) complex dissolved in water trapped inside sephadex particles, suspended in 1-butanol.

Lipase-catalysed acylation of starch and determination of the degree of substitution by methanolysis and GC.

Background: Natural polysaccharides such as starch are becoming increasingly interesting as renewable starting materials for the synthesis of biodegradable polymers using chemical or enzymatic methods. Given the complexity of polysaccharides, the analysis of reaction products is challenging.

Results: Esterification of starch with fatty acids has traditionally been monitored by saponification and back-titration, but in our experience this method is unreliable.

Biocompatible functionalisation of starch.

The enzyme catalysed esterification of starch and fatty acids with terminal triple bonds is described. This material can be used as an acceptor for azide containing molecules, through azide/alkyne cycloaddition. The potential is illustrated by the production of fluorescently-labelled starch, and a biotinylated derivative which can bind streptavidin.