Oxidation states of the manganese cluster during the flash-induced S-state cycle of the photosynthetic oxygen-evolving complex.

The Mn K-edge x-ray absorption spectra for the pure S states of the tetranuclear Mn cluster of the oxygen-evolving complex of photosystem II during flash-induced S-state cycling have been determined. The relative S-state populations in samples given 0, 1, 2, 3, 4, or 5 flashes were determined from fitting the flash-induced electron paramagnetic resonance (EPR) multiline signal oscillation pattern to the Kok model. The edge spectra of samples given 0, 1, 2, or 3 flashes were combined with EPR information to calculate the pure S-state edge spectra. The edge positions (defined as the zero-crossing of the second derivatives) are 6550.1, 6551.7, 6553.5, and 6553.8 eV for S0, S1, S2, and S3, respectively. In addition to the shift in edge position, the S0--> S1 and S1--> S2 transitions are accompanied by characteristic changes in the shape of the edge, both indicative of Mn oxidation. The edge position shifts very little (0.3 eV) for the S2--> S3 transition, and the edge shape shows only subtle changes. We conclude that probably no direct Mn oxidation is involved in this transition. The proposed Mn oxidation state assignments are as follows: S0 (II, III, IV, IV) or (III, III, III, IV), S1 (III, III, IV, IV), S2 (III, IV, IV, IV), S3 (III, IV, IV, IV).

Twenty-five coregulated transcripts define a sterigmatocystin gene cluster in Aspergillus nidulans.

Sterigmatocystin (ST) and the aflatoxins (AFs), related fungal secondary metabolites, are among the most toxic, mutagenic, and carcinogenic natural products known. The ST biosynthetic pathway in Aspergillus nidulans is estimated to involve at least 15 enzymatic activities, while certain Aspergillus parasiticus, Aspergillus flavus, and Aspergillus nomius strains contain additional activities that convert ST to AF. We have characterized a 60-kb region in the A. nidulans genome and find it contains many, if not all, of the genes needed for ST biosynthesis. This region includes verA, a structural gene previously shown to be required for ST biosynthesis, and 24 additional closely spaced transcripts ranging in size from 0.6 to 7.2 kb that are coordinately induced only under ST-producing conditions. Each end of this gene cluster is demarcated by transcripts that are expressed under both ST-inducing and non-ST-inducing conditions. Deduced polypeptide sequences of regions within this cluster had a high percentage of identity with enzymes that have activities predicted for ST/AF biosynthesis, including a polyketide synthase, a fatty acid synthase (alpha and beta subunits), five monooxygenases, four dehydrogenases, an esterase, an 0-methyltransferase, a reductase, an oxidase, and a zinc cluster DNA binding protein. A revised system for naming the genes of the ST pathway is presented.

Z-DNA-forming sites within the human beta-globin gene cluster.

Agarose-encapsulated, metabolically active, permeabilized nuclei from human hematopoietic cell lines were tested for Z-DNA formation in the beta-globin gene cluster. Biotinylated monoclonal antibodies against Z-DNA were diffused into the nuclei and cross-linked to DNA with a 10-ns laser exposure at 266 nm. Following digestion with restriction enzymes, fragments that had formed Z-DNA were isolated. Seventeen regions with Z-DNA sequence motifs in the 73-kb region were studied by PCR amplification, and five were found in the Z conformation.

Site-directed mutagenesis of Azotobacter vinelandii ferredoxin I: cysteine ligation of the [4Fe-4S] cluster with protein rearrangement is preferred over serine ligation.

The [4Fe-4S] cluster of Azotobacter vinelandii ferredoxin I receives three of its four ligands from a Cys-Xaa-Xaa-Cys-Xaa-Xaa-Cys sequence at positions 39-45 while the fourth ligand, Cys20, is provided by a distal portion of the sequence. Previously we reported that the site-directed mutation of Cys20 to Ala (C20A protein) resulted in the formation of a new [4Fe-4S] cluster that obtained its fourth ligand from Cys24, a free cysteine in the native structure. That ligand exchange required significant protein rearrangement. Here we report the conversion of Cys20 to Ser (C20S protein), which gives the protein the opportunity either to retain the native structure and use the Ser20 O gamma as a ligand or to rearrange and use Cys24. X-ray crystallography demonstrates that the cluster does not use the Ser20 O gamma as a ligand; rather it rearranges to use Cys24. In the C20S protein the [4Fe-4S] cluster has altered stability and redox properties relative to either C20A or the native protein.

Role of aromatic residues in stabilization of the [Fe4S4] cluster in high-potential iron proteins (HiPIPs): physical characterization and stability studies of Tyr-19 mutants of Chromatium vinosum HiPIP.

The functional role of residue Tyr-19 of Chromatium vinosum HiPIP has been evaluated by site-directed mutagenesis experiments. The stability of the [Fe4S4] cluster prosthetic center is sensitive to side-chain replacements. Polar residues result in significant instability, while nonpolar residues (especially with aromatic side chains) maintain cluster stability. Two-dimensional NMR data of native and mutant HiPIPs are consistent with a model where Tyr-19 serves to preserve the structural rigidity of the polypeptide backbone, thereby maintaining a hydrophobic barrier for exclusion of water from the cluster cavity. Solvent accessibility results in more facile oxidation of the cluster by atmospheric oxygen, with subsequent rapid hydrolysis of the [Fe4S4]3+ core.

Proximity of the manganese cluster of photosystem II to the redox-active tyrosine YZ.

Electron spin echo electron-nuclear double resonance (ESE-ENDOR) experiments performed on a broad radical electron paramagnetic resonance (EPR) signal observed in photosystem II particles depleted of Ca2+ indicate that this signal arises from the redox-active tyrosine YZ. The tyrosine EPR signal width is increased relative to that observed in a manganese-depleted preparation due to a magnetic interaction between the photosystem II manganese cluster and the tyrosine radical. The manganese cluster is located asymmetrically with respect to the symmetry-related tyrosines YZ and YD. The distance between the YZ tyrosine and the manganese cluster is estimated to be approximately 4.5 A. Due to this close proximity of the Mn cluster and the redox-active tyrosine YZ, we propose that this tyrosine abstracts protons from substrate water bound to the Mn cluster.

Hydrophobic cluster analysis predicts an amino-terminal domain of varicella-zoster virus open reading frame 10 required for transcriptional activation.

Varicella-zoster virus open reading frame 10 (ORF10) protein, the homolog of the herpes simplex virus protein VP16, can transactivate immediate-early promoters from both viruses. A protein sequence comparison procedure termed hydrophobic cluster analysis was used to identify a motif centered at Phe-28, near the amino terminus of ORF10, that strongly resembles the sequence of the activating domain surrounding Phe-442 of VP16. With a series of GAL4-ORF10 fusion proteins, we mapped the ORF10 transcriptional-activation domain to the amino-terminal region (aa 5-79). Extensive mutagenesis of Phe-28 in GAL4-ORF10 fusion proteins demonstrated the importance of an aromatic or bulky hydrophobic amino acid at this position, as shown previously for Phe-442 of VP16. Transactivation by the native ORF10 protein was abolished when Phe-28 was replaced by Ala. Similar amino-terminal domains were identified in the VP16 homologs of other alphaherpesviruses. Hydrophobic cluster analysis correctly predicted activation domains of ORF10 and VP16 that share critical characteristics of a distinctive subclass of acidic activation domains.

The biosynthetic gene cluster for the polyketide immunosuppressant rapamycin.

The macrocyclic polyketides rapamycin and FK506 are potent immunosuppressants that prevent T-cell proliferation through specific binding to intracellular protein receptors (immunophilins). The cloning and specific alteration of the biosynthetic genes for these polyketides might allow the biosynthesis of clinically valuable analogues. We report here that three clustered polyketide synthase genes responsible for rapamycin biosynthesis in Streptomyces hygroscopicus together encode 14 homologous sets of enzyme activities (modules), each catalyzing a specific round of chain elongation. An adjacent gene encodes a pipecolate-incorporating enzyme, which completes the macrocycle. The total of 70 constituent active sites makes this the most complex multienzyme system identified so far. The DNA region sequenced (107.3 kbp) contains 24 additional open reading frames, some of which code for proteins governing other key steps in rapamycin biosynthesis.

The penicillin gene cluster is amplified in tandem repeats linked by conserved hexanucleotide sequences.

The penicillin biosynthetic genes (pcbAB, pcbC, penDE) of Penicillium chrysogenum AS-P-78 were located in a 106.5-kb DNA region that is amplified in tandem repeats (five or six copies) linked by conserved TTTACA sequences. The wild-type strains P. chrysogenum NRRL 1951 and Penicillium notatum ATCC 9478 (Fleming's isolate) contain a single copy of the 106.5-kb region. This region was bordered by the same TTTACA hexanucleotide found between tandem repeats in strain AS-P-78. A penicillin overproducer strain, P. chrysogenum E1, contains a large number of copies in tandem of a 57.9-kb DNA fragment, linked by the same hexanucleotide or its reverse complementary TGTAAA sequence. The deletion mutant P. chrysogenum npe10 showed a deletion of 57.9 kb that corresponds exactly to the DNA fragment that is amplified in E1. The conserved hexanucleotide sequence was reconstituted at the deletion site. The amplification has occurred within a single chromosome (chromosome I). The tandem reiteration and deletion appear to arise by mutation-induced site-specific recombination at the conserved hexanucleotide sequences.

Hα emission fluxes and lithium abundances of low-mass stars in the young open cluster IC 4665

As part of a long term effort to understand pre-main sequence Li burning, we have obtained high resolution spectroscopic observations of 14 late type stars (G0-M1) in the young open cluster IC 4665. Most of the stars have Hα filled-in and Li I absorption, as expected for their young age. From the equivalent widths of Hα emission excess (obtained using the spectral subtraction technique) and the the Li i λ6708 feature, we have derived Hα emission fluxes and photospheric Li abundances. The mean Li abundance of IC 4665 solar-type stars is log N(Li) = 3.1; the same as in other young clusters (α Per, Pleiades) and T Tauri stars. Our results support the conclusions from previous works that PMS Li depletion is very small for masses ∼ 1 M_⨀ . Among the IC 4665 late-G and early K-type stars, there is a spread in Li abundances of about one order of magnitude. The Li-poor IC 4665 members have low Hα excess and vsini≤10. Hence, the Li-activity-rotation connection which has been clearly established in the Pleiades also seems to hold in IC 4665. One M-type IC 4665 star that we have observed does not show Li, implying a very efficient Li depletion as observed in α Per stars of the same spectral type. The level of chromospheric activity and Li depletion among the low-mass stars of IC 4665 is similar to that in the Pleiades. In fact, we note that the Li abundance distributions in several young clusters (α Per, Pleiades, IC 2391, IC 4665) and in post T Tauri stars are strikingly similar. This result suggests that Hα emission and Li abundance not well correlated with age for low-mass stars between 20 and 100 Myr old. We argue that a finer age indicator, the ''LL-clock'', would be the luminosity at which the transition between efficient Li depletion and preservation takes place for fully convective objects. The LL-clock could allow in the near future to derive the relative ages of young open clusters, and clarify the study of PMS evolution of cool stars.

Stars and brown dwarfs in the σ Orionis cluster (Research Note) III. OSIRIS/GTC low-resolution spectroscopy of variable sources

Context. Although many studies have been performed so far, there are still dozens of low-mass stars and brown dwarfs in the young σ Orionis open cluster without detailed spectroscopic characterisation. Aims. We look for unknown strong accretors and disc hosts that were undetected in previous surveys. Methods. We collected low-resolution spectroscopy (R ~ 700) of ten low-mass stars and brown dwarfs in σ Orionis with OSIRIS at the Gran Telescopio Canarias under very poor weather conditions. These objects display variability in the optical, infrared, Hα, and/or X-rays on time scales of hours to years. We complemented our spectra with optical and near-/mid-infrared photometry. Results. For seven targets, we detected lithium in absorption, identified Hα, the calcium doublet, and forbidden lines in emission, and/or determined spectral types for the first time. We characterise in detail a faint, T Tauri-like brown dwarf with an 18 h-period variability in the optical and a large Hα equivalent width of –125  ±  15 Å, as well as two M1-type, X-ray-flaring, low-mass stars, one with a warm disc and forbidden emission lines, the other with a previously unknown cold disc with a large inner hole. Conclusions. New unrevealed strong accretors and disc hosts, even below the substellar limit, await discovery among the list of known σ Orionis stars and brown dwarfs that are variable in the optical and have no detailed spectroscopic characterisation yet.

Deep observation of the NGC1275 region with MAGIC: search of diffuse gamma-ray emission from cosmic rays in the Perseus cluster

Clusters of galaxies are expected to be reservoirs of cosmic rays (CRs) that should produce diffuse γ-ray emission due to their hadronic interactions with the intra-cluster medium. The nearby Perseus cool-core cluster, identified as the most promising target to search for such an emission, has been observed with the MAGIC telescopes at very-high energies (VHE, E ≥ 100 GeV) for a total of 253 hr from 2009 to 2014. The active nuclei of NGC 1275, the central dominant galaxy of the cluster, and IC 310, lying at about 0.6º from the centre, have been detected as point-like VHE γ-ray emitters during the first phase of this campaign. We report an updated measurement of the NGC 1275 spectrum, which is described well by a power law with a photon index Γ = 3.6 ± 0.2_(stat) ± 0.2_(syst) between 90 GeV and 1200 GeV. We do not detect any diffuse γ-ray emission from the cluster and so set stringent constraints on its CR population. To bracket the uncertainties over the CR spatial and spectral distributions, we adopt different spatial templates and power-law spectral indexes α. For α = 2.2, the CR-to-thermal pressure within the cluster virial radius is constrained to be ≤ 1 − 2%, except if CRs can propagate out of the cluster core, generating a flatter radial distribution and releasing the CR-to-thermal pressure constraint to ≤ 20%. Assuming that the observed radio mini-halo of Perseus is generated by secondary electrons from CR hadronic interactions, we can derive lower limits on the central magnetic field, B_(0), that depend on the CR distribution. For α = 2.2, B_(0) ≥ 5 − 8 µG, which is below the ∼25 µG inferred from Faraday rotation measurements, whereas for α ≤ 2.1, the hadronic interpretation of the diffuse radio emission contrasts with our γ-ray flux upper limits independently of the magnetic field strength.

Cluster tecnológicos como fórmula para la renovación de los destinos: hacia un turismo sostenible

En el actual marco de globalización, el análisis de la competitividad turística es cada vez más importante para los destinos, que se enfrentan a un mayor número de competidores y a una sociedad cambiante en relación a gustos, motivos del viaje y formas de moverse. Ante esta perspectiva de cambio, la innovación parece haberse convertido en uno de los principales elementos para la dotación de ventajas competitivas y diferenciación entre rivales. Paralelamente, el análisis de los cluster turísticos ha derivado que de su formación se obtienen incrementos muy importantes en materia de innovación, gracias a la relación competencia-colaboración que tiene lugar entre empresas. Pero hasta la fecha, en el turismo esta relación ha tenido lugar en un marco de cluster industrial, relegando a un segundo plano la innovación como elemento competitivo frente a otra serie de dotaciones. En este documento sin embargo se expone la necesidad de generar clusters basados en la innovación como principal fórmula para evolucionar de acuerdo a las necesidades de la demanda y a las características del mercado, situando a los destinos en una situación de vanguardia y sostenibilidad. El papel de las Universidades y del capital humano serán fundamentales para la adaptación del turismo en una industria centrada en la innovación.

An accurate distance scale to the extremely massive cluster Stephenson 2

Discoveries during the last two years have revealed the existence of a vast region of star formation close to the base of the Scutum Arm, where at least five clusters of red supergiants have been found. In order to understand the nature of this region, we need to determine accurate distances to the clusters. We present here the first results of an ongoing program to derive fundamental parameters (such as age, distance, etc.) to the massive cluster Stephenson 2 studying for the first time its main sequence stars.