Studies of viomycin, an anti-tuberculosis antibiotic: Copper(II) coordination, DNA degradation and the impact on delta ribozyme cleavage activity

Viomycin is a basic peptide antibiotic, which is among the most effective agents against multidrug-resistant tuberculosis. In this paper we provide the characteristics of its acid base properties, coordination preferences towards the Cu(II) ions, as well as the reactivity of the resulting complexes against plasmid DNA and HDV ribozyme. Careful coordination studies throughout the wide pH range allow for the characterisation of all the Cu(II)-viomycin complex species. The assignment of proton chemical shifts was achieved by NMR experiments, while the DTF level of theory was applied to support molecular structures of the studied complexes. The experiments with the plasmid DNA reveal that at the physiological levels of hydrogen peroxide the Cu(II)-viomycin complex is more aggressive against DNA than uncomplexed metal ions. Moreover, the degradation of DNA by viomycin can be carried out without the presence of transition metal ions. In the studies of antigenomic delta ribozyme catalytic activity, viomycin and its complex are shown to modulate the ribozyme functioning. The molecular modelling approach allows the indication of two different locations of viomycin binding sites to the ribozyme

The Myxococcus xanthus Two-Component System CorSR Regulates Expression of a Gene Cluster Involved in Maintaining Copper Tolerance during Growth and Development

Myxococcus xanthus is a soil-dwelling member of the δ–Proteobacteria that exhibits a complex developmental cycle upon starvation. Development comprises aggregation and differentiation into environmentally resistant myxospores in an environment that includes fluctuations in metal ion concentrations. While copper is essential for M. xanthus cells because several housekeeping enzymes use it as a cofactor, high copper concentrations are toxic. These opposing effects force cells to maintain a tight copper homeostasis. A plethora of paralogous genes involved in copper detoxification, all of which are differentially regulated, have been reported in M. xanthus. The use of in-frame deletion mutants and fusions with the reporter gene lacZ has allowed the identification of a two-component system, CorSR, that modulates the expression of an operon termed curA consisting of nine genes whose expression slowly increases after metal addition, reaching a plateau. Transcriptional regulation of this operon is complex because transcription can be initiated at different promoters and by different types of regulators. These genes confer copper tolerance during growth and development. Copper induces carotenoid production in a ΔcorSR mutant at lower concentrations than with the wild-type strain due to lack of expression of a gene product resembling subunit III of cbb3-type cytochrome c oxidase. This data may explain why copper induces carotenoid biosynthesis at suboptimal rather than optimal growth conditions in wild-type strains.This work has been funded by the Spanish Government (grants CSD2009-00006 and BFU2012-33248, 70% funded by FEDER). This work was also supported by the National Institute of General Medical Science of the National Institutes of Health under award number R01GM095826 to LJS, and by the National Science Foundation under award number MCB0742976 to LJS. JMD and JP received a fellowship from Junta de Andalucía to do some work at University of Georgia

Positive Evolutionary Selection of an HD Motif on Alzheimer Precursor Protein Orthologues Suggests a Functional Role

HD amino acid duplex has been found in the active center of many different enzymes. The dyad plays remarkably different roles in their catalytic processes that usually involve metal coordination. An HD motif is positioned directly on the amyloid beta fragment (Aβ) and on the carboxy-terminal region of the extracellular domain (CAED) of the human amyloid precursor protein (APP) and a taxonomically well defined group of APP orthologues (APPOs). In human Aβ HD is part of a presumed, RGD-like integrin-binding motif RHD; however, neither RHD nor RXD demonstrates reasonable conservation in APPOs. The sequences of CAEDs and the position of the HD are not particularly conserved either, yet we show with a novel statistical method using evolutionary modeling that the presence of HD on CAEDs cannot be the result of neutral evolutionary forces (p<0.0001). The motif is positively selected along the evolutionary process in the majority of APPOs, despite the fact that HD motif is underrepresented in the proteomes of all species of the animal kingdom. Position migration can be explained by high probability occurrence of multiple copies of HD on intermediate sequences, from which only one is kept by selective evolutionary forces, in a similar way as in the case of the “transcription binding site turnover.” CAED of all APP orthologues and homologues are predicted to bind metal ions including Amyloid-like protein 1 (APLP1) and Amyloid-like protein 2 (APLP2). Our results suggest that HDs on the CAEDs are most probably key components of metal-binding domains, which facilitate and/or regulate inter- or intra-molecular interactions in a metal ion-dependent or metal ion concentration-dependent manner. The involvement of naturally occurring mutations of HD (Tottori (D7N) and English (H6R) mutations) in early onset Alzheimer's disease gives additional support to our finding that HD has an evolutionary preserved function on APPOs

CMS physics technical design report : Addendum on high density QCD with heavy ions

Peer reviewe

Iron Behaving Badly: Inappropriate Iron Chelation as a Major Contributor to the Aetiology of Vascular and Other Progressive Inflammatory and Degenerative Diseases

The production of peroxide and superoxide is an inevitable consequence of aerobic metabolism, and while these particular "reactive oxygen species" (ROSs) can exhibit a number of biological effects, they are not of themselves excessively reactive and thus they are not especially damaging at physiological concentrations. However, their reactions with poorly liganded iron species can lead to the catalytic production of the very reactive and dangerous hydroxyl radical, which is exceptionally damaging, and a major cause of chronic inflammation. We review the considerable and wide-ranging evidence for the involvement of this combination of (su)peroxide and poorly liganded iron in a large number of physiological and indeed pathological processes and inflammatory disorders, especially those involving the progressive degradation of cellular and organismal performance. These diseases share a great many similarities and thus might be considered to have a common cause (i.e. iron-catalysed free radical and especially hydroxyl radical generation). The studies reviewed include those focused on a series of cardiovascular, metabolic and neurological diseases, where iron can be found at the sites of plaques and lesions, as well as studies showing the significance of iron to aging and longevity. The effective chelation of iron by natural or synthetic ligands is thus of major physiological (and potentially therapeutic) importance. As systems properties, we need to recognise that physiological observables have multiple molecular causes, and studying them in isolation leads to inconsistent patterns of apparent causality when it is the simultaneous combination of multiple factors that is responsible. This explains, for instance, the decidedly mixed effects of antioxidants that have been observed, etc...Comment: 159 pages, including 9 Figs and 2184 reference

Metal ion effects on cis/trans isomerization of proline residues in short chain peptides in solution

The-effect of capper(II) ions on the probabilities of existence of the four detectable conformers of the tetrapeptide Tyr-Pro-Phe-Pro (beta -casomorphin 4) in [H-2(6)]DMSO was investigated by H-1 NMR Spectroscopy. Integration of the Phe-NH signals provided the relative populations in the free state as tt/tc/ct/cc = 28.34.29:9 at 293 K (c = cis, t = trans). Copper(ii) was shown to bind to all four isomers, yielding complexes with two different structures, depending on the conformation of Pro(2). The interpretation of paramagnetic relaxation rates of Pro(2)-H alpha signals provided the corresponding isomeric probabilities in the metal-bound state as 13:36:20:31. The observed stabilization of the conformation with the lowest probability of-existence (cc) may be relevant for the biological role of copper and other metal ions

Calcium ions affect the exchange network but not the structure of a small peptide (melanostatin) in solution: a 1H- and 13C-NMR study

The interaction of calcium ions with the peptide hormone melanostatin (Pro-Leu-Gly-NH2) was investigated by H-1 and C-13 NMR spectroscopy in [D-6]DMSO containing H2O (1%). Chemical shifts, spin-lattice relaxation rates, H-1 NOESY maps and the temperature coefficients of the amide 1H NMR chemical shifts were measured at increasing concentrations of calcium. A 1:1 complex with the metal coordinated to the carbonyl moieties of Pro and Gly (K-d = 17 +/- 2 mM(-1)) was shown to be the major species in solution, although evidence was also provided for the occurrence of a minor species with the metal bound to the Leu carbonyl and with different stoichiometry. Upon metal complexation, substantial changes in the intrinsic chain flexibility of the peptide and in the exchange rates between water and amide protons were detected

Structural features of apramycin bound at the bacterial ribosome a site as detected by NMR and CD spectroscopy

The interaction of apramycin with an RNA fragment mimicking the bacterial ribosomal A site was investigated by NMR spectroscopy, mainly by using the transferred NOE technique. The fact that only the RNA-bound antibiotic shows nonzero NOE effects, allowed us to gain structural details of bound apramycin despite the fast exchange conditions between the free and RNA-bound forms. The dissociation constant of the apramycin–RNA complex was evaluated by CD spectroscopy

Interaction of Angiotensin II with the C-terminal 300-320 fragment of the rat Angiotensin II Receptor AT1a monitored by NMR.

Interaction between angiotensin II (Ang II) and the fragment peptide 300–320 (fCT300–320) of the rat angiotensin II receptor AT1a was demonstrated by relaxation measurements, NOE effects, chemical shift variations, and CD measurements. The correlation times modulating dipolar interactions for the bound and free forms of Ang II were estimated by the ratio of the nonselective and single-selective longitudinal relaxation rates. The intermolecular NOEs observed in NOESY spectra between HN protons of 9LysfCT and 6Hisang, 10PhefCT and 8Pheang, HN proton of 3TyrfCT and Hα of 4Tyrang, 5PhefCTHδ and Hα of 4Tyrang indicated that Ang II aromatic residues are directly involved in the interaction, as also verified by relaxation data. Some fCT300–320 backbone features were inferred by the CSI method and CD experiments revealing that the presence of Ang II enhances the existential probability of helical conformations in the fCT fragment. Restrained molecular dynamics using the simulated annealing protocol was performed with intermolecular NOEs as constraints, imposing an α-helix backbone structure to fCT300–320 fragment. In the built model, one strongly preferred interaction was found that allows intermolecular stacking between aromatic rings and forces the peptide to wrap around the 6Leu side chain of the receptor fragment