Stabilisation of tyrosinase by reversed micelles for bioelectrocatalysis in dry organic media

The enzymatic and bioelectrocatalytic activity of tyrosinase from mushrooms was studied in a system of reversed micelles formed by Aerosol OT (AOT) in hexane. The optimal catechol oxidising activity of tyrosinase incorporated in reversed micelles was found at a hydration degree of w(0) = 25. The catalytic activity was comparable with tyrosinase activity in aqueous media. When immobilized at an Au electrode, either directly or in reversed micelles, tyrosinase exhibited a similar efficiency of the bioelectrocatalytic reduction of O-2 mediated by catechol; however, a rapid decrease in the activity correlated with the destruction of reversed micelles and/or the removal of tyrosinase from the electrode surface. The system containing tyrosinase in reversed micelles with caoutchouk, spread on the surface of the An electrode and successively covered with a Nafion(R) membrane layer, was found to result in stable tyrosinase-modified electrodes, which were resistant to inactivation in dry acetonitrile. The proposed technique offers possibilities for further development of highly active and stable surfactant/enzyme-modified electrodes for measurements carried out in organic solvents. (C) 2002 Elsevier Science B.V. All rights reserved

Electrocatalytic interconversion of NADH and NAD+ by Escherichia coli flavohemoglobin

E. coli flavohemoglobin, oriented at electrodes via amphiphilic polymyxin B, electrocatalytically interconverts NADH and NAD(+) at its heme potentials operating as an electron transfer relay between the electrode and the protein FAD, where NADH/NAD(+) is transformed. The results are crucial for the development of NAD(+)-dependent bioelectrodes for biosynthesis, biosensors and biofuel cells. PMID: 26389555 [PubMed - in process

Water-in-Oil Micro-Emulsion Enhances the Secondary Structure of a Protein by Confinement

A scheme is presented in which an organic solvent environment in combination with surfactants is used to confine a natively unfolded protein inside an inverse microemulsion droplet. This type of confinement allows a study that provides unique insight into the dynamic structure of an unfolded, flexible protein which is still solvated and thus under near-physiological conditions. In a model system, the protein osteopontin (OPN) is used. It is a highly phosphorylated glycoprotein that is expressed in a wide range of cells and tissues for which limited structural analysis exists due to the high degree of flexibility and large number of post-translational modifications. OPN is implicated in tissue functions, such as inflammation and mineralisation. It also has a key function in tumour metastasis and progression. Circular dichroism measurements show that confinement enhances the secondary structural features of the protein. Small-angle X-ray scattering and dynamic light scattering show that OPN changes from being a flexible protein in aqueous solution to adopting a less flexible and more compact structure inside the microemulsion droplets. This novel approach for confining proteins while they are still hydrated may aid in studying the structure of a wide range of natively unfolded proteins.Danish National Advanced Technology Foundation through the ProSURF platform (Protein-Based Functionalisation of Surfaces)Danish National Advanced Technology Foundation through the ProSURF platform (ProteinBased Functionalisation of Surfaces)iNANO center from the Danish Research CouncilsiNANO center from the Danish Research CouncilsVillum Kahn Rasmussen FoundationVillum Kahn Rasmussen FoundationLundbeck FoundationLundbeck FoundationCarlsberg FoundationCarlsberg Foundatio