Anomalous x-ray scattering from terbium-labeled parvalbumin in solution

We have used anomalous small-angle x-ray scattering as a structural probe for solutions of rabbit parvalbumin labeled with terbium. This technique makes use of the large changes in the terbium scattering factor that occur when the x-ray energy is tuned around an L3 absorption edge of this heavy-atom label. These changes in scattering result in changes in the small-angle scattering curve of the labeled protein as a whole, which can then be analyzed to derive structural information concerning the distribution of labels in the protein. Based on a Gaussian model for the protein electron density, the mean distance from the terbiums to the protein center of mass is determined to be 13.2 A and is consistent with crystallographic results. Our results demonstrate the usefulness of terbium as an anomalous scattering label and provide criteria to help establish anomalous scattering as a reliable structural technique for proteins in solution

PySpline: A Modern, Cross-Platform Program for the Processing of Raw Averaged XAS Edge and EXAFS Data

PySpline is a modern computer program for processing raw averaged XAS and EXAFS data using an intuitive approach which allows the user to see the immediate effect of various processing parameters on the resulting k- and R-space data. The Python scripting language and Qt and Qwt widget libraries were chosen to meet the design requirement that it be cross-platform (i.e. versions for Windows, Mac OS X, and Linux). PySpline supports polynomial pre- and post-edge background subtraction, splining of the EXAFS region with a multi-segment polynomial spline, and Fast Fourier Transform (FFT) of the resulting k{sup 3}-weighted EXAFS data

Gene expression patterns unveil a new level of molecular heterogeneity in colorectal cancer.

The recognition that colorectal cancer (CRC) is a heterogeneous disease in terms of clinical behaviour and response to therapy translates into an urgent need for robust molecular disease subclassifiers that can explain this heterogeneity beyond current parameters (MSI, KRAS, BRAF). Attempts to fill this gap are emerging. The Cancer Genome Atlas (TGCA) reported two main CRC groups, based on the incidence and spectrum of mutated genes, and another paper reported an EMT expression signature defined subgroup. We performed a prior free analysis of CRC heterogeneity on 1113 CRC gene expression profiles and confronted our findings to established molecular determinants and clinical, histopathological and survival data. Unsupervised clustering based on gene modules allowed us to distinguish at least five different gene expression CRC subtypes, which we call surface crypt-like, lower crypt-like, CIMP-H-like, mesenchymal and mixed. A gene set enrichment analysis combined with literature search of gene module members identified distinct biological motifs in different subtypes. The subtypes, which were not derived based on outcome, nonetheless showed differences in prognosis. Known gene copy number variations and mutations in key cancer-associated genes differed between subtypes, but the subtypes provided molecular information beyond that contained in these variables. Morphological features significantly differed between subtypes. The objective existence of the subtypes and their clinical and molecular characteristics were validated in an independent set of 720 CRC expression profiles. Our subtypes provide a novel perspective on the heterogeneity of CRC. The proposed subtypes should be further explored retrospectively on existing clinical trial datasets and, when sufficiently robust, be prospectively assessed for clinical relevance in terms of prognosis and treatment response predictive capacity. Original microarray data were uploaded to the ArrayExpress database (http://www.ebi.ac.uk/arrayexpress/) under Accession Nos E-MTAB-990 and E-MTAB-1026. © 2013 Swiss Institute of Bioinformatics. Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland

New Developments in Charge Transfer Multiplet Calculations: Projection Operations, Mixed-Spin States and pi-Bonding

This paper presents a number of new additions to the charge transfer multiplet calculations as used in the calculation of L edge X-ray absorption spectra of 3d and 4d transition metal systems, both oxides and coordination compounds. The focus of the paper is on the consequences of the optimized spectral simulations for the ground state, where we make use of a recently developed projection technique. This method is also used to develop the concept of a mixed-spin ground state, i.e. a state that is a mixture of a high-spin and low-spin state due to spin-orbit coupling combined with strong covalency. The charge transfer mechanism to describe {pi}-bonding uses the mixing of the metal-to-ligand charge transfer (MLCT) channel in addition to the normal CT channel and allows for the accurate simulation of {pi}-bonding systems, for example cyanides

XANES/EXAFS STUDY OF THE COPPER ACTIVE SITE IN METHANOL SYNTHESIS CATALYST

We have used X-ray absorption spectroscopy to determine the electronic and geometric structure of the copper active site in methanol synthesis catalysts. Compared to the edge and FT of the EXAFS data of CuO, the data suggest that most of the Cu(II) in the calcined form does not have the same ligation as in the CuO lattice and more importantly that some of the Cu is dissolved in the ZnO lattice. For the reduced sample, the edge shifts to lower energy after reduction, indicating the presence of either Cu(I) or Cu(0). The EXAFS data analyses of reduced catalysts show the presence of copper clusters. Reaction with H2S is shown to decompose the Cu clusters in the reduced catalyst and the level of Cu cluster decomposition increases with further reaction at higher temperature and longer time period

EXAFS STUDY OF ACTIVE INTERMEDIATES : HEME ENZYMES AND MODEL COMPOUNDS

The iron coordination structures of the oxygenated cytochrome P-450-CAM and chloroperoxidase and of a number of penta- and hexa-coordinate ferrous porphyrin complexes containing biologically relevant, sulfur axial ligands have been investigatied by EXAFS spectroscopy. The two oxygenated enzymes have both been found to contain a sulfur atom located 2.37 Å from the central heme iron. In the CO-ligated series of model complexes, two distinct categories of Fe-Sax distances have been observed that correlate with thiolate and non-thiolate ligation (~2.33 and ~2.41 Å, respectively) . The data for these model ferrous-CO ccomplexes, together with previously reported Fe-S distance in oxygenated heme iron model complexes ; allows the sulfur atoms in oxygenated P-450-CAM and chloroperoxidase to be identified as a thiolate sulfur