Prediction of the mechanisms of enzyme-catalysed reactions using hybrid quantum mechanical/molecular mechanical methods

The use of hybrid methods, involving both quantum mechanics and molecular mechanics, to model the mechanism of enzyme-catalysed reactions, is discussed. Two alternative approaches to treating the electrostatic interactions between the quantum mechanical and molecular mechanical regions are studied, involving either the inclusion of this term in the electronic Hamiltonian (QM/MM), or evaluating it purely classically (MO + MM). In the latter scheme, possible problems of using force fields that are standard for macromolecular modelling are identified. The use of QM/MM schemes to investigate the mechanism of the enzymes thymidine phosphorylase (ThdPase) and protein tyrosine phosphatase (PTP) is described. For both systems, transition states have been identified using a PM3 Hamiltonian. For ThdPase, concerted motion of the enzyme during the course of the reaction is suggested and, for PTP, a two-step dephosphorylation reaction is indicated, both with quite low barriers.     

A comparison of lobe and cartesian gaussian basis sets for molecular calculations

Lobe gaussian and cartesian gaussian basis sets, of approximately minimal basis Slater accuracy, have been compared for molecular calculations. The basis sets were constructed so that they only differed in the representation of the angular dependence of the p function. Calculation of total energy and several one-electron properties for a series of nine molecules shows that, for molecular calculations, the lobe and cartesian gaussian representations are equivalent.     

3D reconstruction of the brain from magnetic resonance images using a connectivity algorithm

We present high resolution three dimensional (3D) connectivity, surface construction and display algorithms that detect, extract, and display the surface of a brain from contiguous magnetic resonance (MR) images. The algorithms identify the external brain surface and create a 3D image, showing the fissures and surface convolutions of the cerebral hemispheres, cerebellum, and brain stem. Images produced by these algorithms also show the morphology of other soft tissue boundaries such as the cerebral ventricular system and the skin of the patient. For the purposes of 3D reconstruction, our experiments show that T1 weighted images give better contrast between the surface of the brain and the cerebral spinal fluid than T2 weighted images. 3D reconstruction of MR data provides a non-invasive procedure for examination of the brain surface and other anatomical features.     

Matrix assisted laser desorption ionisation ion mobility separation mass spectrometry imaging of ex-vivo human skin

Matrix assisted laser desorption ionisation ion mobility separation mass spectrometry imaging (MALDI-IMS-MSI) has been employed to image the distribution of proteins in ex-vivo human skin. Using a “bottom-up” proteomics approach employing “on-tissue” digestion the distribution of abundant skin proteins; collagen, keratin, decorin and serum albumin could be mapped. Images have been recorded at 150 and 30 μm spatial resolution. Multivariate statistical analysis of the data has been employed to associate specific proteins with layers of the skin. The improved specificity given by the use of ion mobility separation in mass spectrometric imaging has been demonstrated by separation of peptide ions from phospholipids.     

An assay for the enzyme N-acetyl-beta-D-glucosaminidase (NAGase) based on electrochemical detection using screen-printed carbon electrodes (SPCEs).

An electrochemical assay for the enzyme N-acetyl-beta-D-glucosaminidase (NAGase) is described, using bare screen-printed carbon electrodes (SPCEs). The enzyme substrate, 1-naphthyl-N-acetyl-beta-D-glucosaminide, was added to the NAGase-containing sample under hydrodynamic conditions and was hydrolysed to 1-naphthol, which was monitored amperometrically at an Eapp of +650 mV versus SCE. A pH study revealed the apparent Vmax for the assay to occur at pH 4.5. corresponding to an apparent substrate Km of 0.28 mM. In order to be compatible with the analysis of biological fluids, a final operating pH of 5.4 was selected, and, using a data recording time of 100 s post-substrate addition, the assay gave a linear response (r2 = 0.988) over the range 3.1 to 108 mU ml(-1) NAGase (RSD = 15.4%). This assay has the potential to monitor NAGase levels in a number of application areas.     

A separable non-remainder of

We prove that there is a compact separable continuum that (consistently) is not a remainder of the real line.