The structure of genetically modified iron-sulfur cluster F(x) in photosystem I as determined by X-ray absorption spectroscopy.

Photosystem I (PS I) mediates light-induced electron transfer from P700 through a chlorophyll a, a quinone and a [4Fe-4S] iron-sulfur cluster F(X), located on the core subunits PsaA/B to iron-sulfur clusters F(A/B) on subunit PsaC. Structure function relations in the native and in the mutant (psaB-C565S/D566E) of the cysteine ligand of F(X) cluster were studied by X-ray absorption spectroscopy (EXAFS) and transient spectroscopy. The structure of F(X) was determined in PS I lacking clusters F(A/B) by interruption of the psaC2 gene of PS I in the cyanobacterium Synechocystis sp PCC 6803. PsaC-deficient mutant cells assembled the core subunits of PS I which mediated electron transfer mostly to the phylloquinone. EXAFS analysis of the iron resolved a [4Fe-4S] cluster in the native PsaC-deficient PS I. Each iron had 4 sulfur and 3 iron atoms in the first and second shells with average Fe-S and Fe-Fe distances of 2.27 A and 2.69 A, respectively. In the C565S/D566E serine mutant, one of the irons of the cluster was ligated to three oxygen atoms with Fe-O distance of 1.81 A. The possibility that the structural changes induced an increase in the reorganization energy that consequently decreased the rate of electron transfer from the phylloquinone to F(X) is discussed.

Zinc cysteine active sites of metalloproteins: a density functional theory and x-ray absorption fine structure study.

Density functional theory (DFT) and x-ray absorption fine structure (XAFS) spectroscopy are complementary tools for the biophysical study of active sites in metalloproteins. DFT is used to compute XAFS multiple scattering Debye Waller factors, which are then employed in genetic algorithm-based fitting process to obtain a global fit to the XAFS in the space of fitting parameters. Zn-Cys sites, which serve important functions as transcriptional switches in Zn finger proteins and matrix metalloproteinases, previously have proven intractable by this method; here these limitations are removed. In this work we evaluate optimal DFT nonlocal functionals and basis sets for determining optimal geometries and vibrational densities of states of mixed ligation Zn(His)(4-n)(Cys)(n) sites. Theoretical results are compared to experimental XAFS measurements and Raman spectra from the literature and tabulated for use.

Ab initio self-consistent x-ray absorption fine structure analysis for metalloproteins.

X-ray absorption fine structure is a powerful tool for probing the structures of metals in proteins in both crystalline and noncrystalline environments. Until recently, a fundamental problem in biological XAFS has been that ad hoc assumptions must be made concerning the vibrational properties of the amino acid residues that are coordinated to the metal to fit the data. Here, an automatic procedure for accurate structural determination of active sites of metalloproteins is presented. It is based on direct multiple-scattering simulation of experimental X-ray absorption fine structure spectra combining electron multiple scattering calculations with density functional theory calculations of vibrational modes of amino acid residues and the genetic algorithm differential evolution to determine a global minimum in the space of fitting parameters. Structure determination of the metalloprotein active site is obtained through a self-consistent iterative procedure with only minimal initial information.

A band dispersion mechanism for Pt alloy compositional tuning of linear bound CO stretching frequencies.

The C-O stretching frequency (nu(CO)) of atop CO/Pt in PtRu alloys is compositionally tuned in proportion to the Pt mole percent. The application of a Blyholder-Bagus type mechanism (i.e., increased back-donation from the metal d-band to the hybridized 2pi CO molecular orbitals (MOs)) to compositional tuning has been paradoxical because (1) a Pt-C bond contraction, expected with increased back-donation as the Pt mole percent is reduced, is not observed (i.e., calculated Pt-C bond is either elongated or insensitive to alloying and the binding energies of CO/Pt decrease with alloying) and (2) the lowering d-band center and increased d-band vacancies upon alloying (suggesting less back-donation to the higher energy metal hybridized 2pi CO MOs) must be reconciled with the alloy-induced red shift of the nu(CO). A library of spin-optimized Pt and Pt alloy clusters was the basis of density functional theory (DFT) calculations of CO binding energies, nu(CO) values, shifts, and broadening of 5sigma/2pi CO MO upon hybridization with the alloy orbitals and a DFT derived Mulliken electron population analysis. The DFT results, combined with FEFF8 local density of states (LDOS) calculations, validate a 5sigma donation-2pi back-donation mechanism, reconciling the direction of alloy compositional tuning with the lowering of the d-band center and increased vacancies. Although the d-band center decreases in energy with alloying, an asymmetric increase in the dispersion of the d-band is accompanied by an upshift of the metal cluster HOMO level. Concomitantly, the hybridization and renormalization of the CO 5sigma/2pi states results in a broadening of the 5sigma/2pi manifold with additional lower energy states closer to the upshifted (with respect to the pure Pt cluster) HOMO of the alloy cluster. The dispersion toward higher energies of the alloy d-density of states results in more 5sigma/2pi CO filled states (i.e., enhanced 2pi-back-donation). Finally, Mulliken and FEFF8 electron population analysis shows that the increase of the average d-band vacancies upon alloying and additional 2pi back-donation are not mutually exclusive. The d-electron density of the CO-adsorbed Pt atom increases with alloying while the average d-electron density throughout the cluster is reduced. The localized electron density is manifested as an electrostatic wall effect, preventing the Pt-C bond contractions expected with increased back-donation to the 2pi CO MOs.

New methods for EXAFS analysis in structural genomics.

Data analysis is one of the remaining bottlenecks in high-throughput EXAFS for structural genomics. Here some recent developments in methodology are described that offer the potential for rapid and automated XAS analysis of metalloproteins.

EXAFS study of Zn sorption mechanisms on montmorillonite.

Extended X-ray absorption fine structure (EXAFS) analysis of zinc sorption on montmorillonite showed that different types of surface complexes or surface precipitates were formed depending on the reaction time. With an initial zinc concentration of 10(-3) M at neutral pH, zinc remained octahedrally coordinated with about six oxygen atoms at Zn-O bond distances of 2.02-2.07 A for up to six months. For samples aged up to 11 days, the Zn-Zn contribution in the second shell suggested formation of multinuclear surface complexes or surface precipitates. For samples aged 20 days and more, Zn-Zn and Zn-Si contributions in the second shell suggested formation of mixed metal coprecipitates such as a Zn phyllosilicate-like phase. Formation of these mixed metal solids probably accounts for the slow continuous sorption reaction at aging times exceeding 20 days. Sequestration of Zn in mixed metal precipitates and the stability of these phases can reduce the concentration, mobility, and toxicity of Zn in soils or sediments.

A device for selecting and rejecting X-ray harmonics in synchrotron radiation beams.

A practical device ('beam cleaner') is described that selects a specific harmonic (e.g. third order) from other nearby harmonics (e.g. fourth or fifth orders) that are transmitted by the beamline monochromator. The ability to isolate and use a clean harmonic beam effectively extends the energy range of existing beamlines. This device is a tunable medium-resolution X-ray bandpass filter, which consists of a double bent Laue crystal post-monochromator. It is a simple and inexpensive device that does not require precision goniometry and can be operated in air. The device can also be used to passively compensate for vertical beam motion as the primary beamline monochromator is scanned. Experimental results are presented that demonstrate the improvement in XAFS data quality under high harmonic conditions.