A distinct, high-affinity, alkaline phosphatase facilitates occupation of P-depleted environments by marine picocyanobacteria.

Marine picocyanobacteria of the genera Prochlorococcus and Synechococcus, the two most abundant phototrophs on Earth, thrive in oligotrophic oceanic regions. While it is well known that specific lineages are exquisitely adapted to prevailing in situ light and temperature regimes, much less is known of the molecular machinery required to facilitate occupancy of these low-nutrient environments. Here, we describe a hitherto unknown alkaline phosphatase, Psip1, that has a substantially higher affinity for phosphomonoesters than other well-known phosphatases like PhoA, PhoX, or PhoD and is restricted to clade III Synechococcus and a subset of high light I-adapted Prochlorococcus strains, suggesting niche specificity. We demonstrate that Psip1 has undergone convergent evolution with PhoX, requiring both iron and calcium for activity and likely possessing identical key residues around the active site, despite generally very low sequence homology. Interrogation of metagenomes and transcriptomes from TARA oceans and an Atlantic Meridional transect shows that psip1 is abundant and highly expressed in picocyanobacterial populations from the Mediterranean Sea and north Atlantic gyre, regions well recognized to be phosphorus (P)-deplete. Together, this identifies psip1 as an important oligotrophy-specific gene for P recycling in these organisms. Furthermore, psip1 is not restricted to picocyanobacteria and is abundant and highly transcribed in some α-proteobacteria and eukaryotic algae, suggesting that such a high-affinity phosphatase is important across the microbial taxonomic world to occupy low-P environments.

Coordinated transcriptional response to environmental stress by a Synechococcus virus.

Viruses are a major control on populations of microbes. Often, their virulence is examined in controlled laboratory conditions. Yet, in nature, environmental conditions lead to changes in host physiology and fitness that may impart both costs and benefits on viral success. Phosphorus (P) is a major abiotic control on the marine cyanobacterium Synechococcus. Some viruses infecting Synechococcus have acquired, from their host, a gene encoding a P substrate binding protein (PstS), thought to improve virus replication under phosphate starvation. Yet, pstS is uncommon among cyanobacterial viruses. Thus, we asked how infections with viruses lacking PstS are affected by P scarcity. We show that the production of infectious virus particles of such viruses is reduced in low P conditions. However, this reduction in progeny is not caused by impaired phage genome replication, thought to be a major sink for cellular phosphate. Instead, transcriptomic analysis showed that under low P conditions, a PstS-lacking cyanophage increased the expression of a specific gene set that included mazG, hli2, and gp43 encoding a pyrophosphatase, a high-light inducible protein and DNA polymerase, respectively. Moreover, several of the upregulated genes were controlled by the host's phoBR two-component system. We hypothesize that recycling and polymerization of nucleotides liberates free phosphate and thus allows viral morphogenesis, albeit at lower rates than when phosphate is replete or when phages encode pstS. Altogether, our data show how phage genomes, lacking obvious P-stress-related genes, have evolved to exploit their host's environmental sensing mechanisms to coordinate their own gene expression in response to resource limitation.

Quantum Dynamics of Oblique Vibrational States in the Hénon-Heiles System.

In this paper, we study the quantum time evolution of oblique nonstationary vibrational states in a Hénon-Heiles oscillator system with two dissociation channels, which models the stretching vibrational motions of triatomic molecules. The oblique nonstationary states we are interested in are the eigenfunctions of the anharmonic zero-order Hamiltonian operator resulting from the transformation to oblique coordinates, which are defined as those coming from nonorthogonal coordinate rotations that express the matrix representation of the second-order Hamiltonian in a block diagonal form characterized by the polyadic quantum number n = n1 + n2. The survival probabilities calculated show that the oblique nonstationary states evolve within their polyadic group with a high degree of coherence up to the dissociation limits on the short time scale. The degree of coherence is certainly much higher than that exhibited by the local nonstationary states extracted from the conventional orthogonal rotation of the original normal coordinates. We also show that energy exchange between the oblique vibrational modes occurs in a much more regular way than the exchange between the local modes.

Bacterial catabolism of membrane phospholipids links marine biogeochemical cycles.

In marine systems, the availability of inorganic phosphate can limit primary production leading to bacterial and phytoplankton utilization of the plethora of organic forms available. Among these are phospholipids that form the lipid bilayer of all cells as well as released extracellular vesicles. However, information on phospholipid degradation is almost nonexistent despite their relevance for biogeochemical cycling. Here, we identify complete catabolic pathways for the degradation of the common phospholipid headgroups phosphocholine (PC) and phosphorylethanolamine (PE) in marine bacteria. Using Phaeobacter sp. MED193 as a model, we provide genetic and biochemical evidence that extracellular hydrolysis of phospholipids liberates the nitrogen-containing substrates ethanolamine and choline. Transporters for ethanolamine (EtoX) and choline (BetT) are ubiquitous and highly expressed in the global ocean throughout the water column, highlighting the importance of phospholipid and especially PE catabolism in situ. Thus, catabolic activation of the ethanolamine and choline degradation pathways, subsequent to phospholipid metabolism, specifically links, and hence unites, the phosphorus, nitrogen, and carbon cycles.

Elucidating the picocyanobacteria salinity divide through ecogenomics of new freshwater isolates.

BACKGROUND: Cyanobacteria are the major prokaryotic primary producers occupying a range of aquatic habitats worldwide that differ in levels of salinity, making them a group of interest to study one of the major unresolved conundrums in aquatic microbiology which is what distinguishes a marine microbe from a freshwater one? We address this question using ecogenomics of a group of picocyanobacteria (cluster 5) that have recently evolved to inhabit geographically disparate salinity niches. Our analysis is made possible by the sequencing of 58 new genomes from freshwater representatives of this group that are presented here, representing a 6-fold increase in the available genomic data. RESULTS: Overall, freshwater strains had larger genomes (≈2.9 Mb) and %GC content (≈64%) compared to brackish (2.69 Mb and 64%) and marine (2.5 Mb and 58.5%) isolates. Genomic novelties/differences across the salinity divide highlighted acidic proteomes and specific salt adaptation pathways in marine isolates (e.g., osmolytes/compatible solutes - glycine betaine/ggp/gpg/gmg clusters and glycerolipids glpK/glpA), while freshwater strains possessed distinct ion/potassium channels, permeases (aquaporin Z), fatty acid desaturases, and more neutral/basic proteomes. Sulfur, nitrogen, phosphorus, carbon (photosynthesis), or stress tolerance metabolism while showing distinct genomic footprints between habitats, e.g., different types of transporters, did not obviously translate into major functionality differences between environments. Brackish microbes show a mixture of marine (salt adaptation pathways) and freshwater features, highlighting their transitional nature. CONCLUSIONS: The plethora of freshwater isolates provided here, in terms of trophic status preference and genetic diversity, exemplifies their ability to colonize ecologically diverse waters across the globe. Moreover, a trend towards larger and more flexible/adaptive genomes in freshwater picocyanobacteria may hint at a wider number of ecological niches in this environment compared to the relatively homogeneous marine system.

Human root dentin microhardness and degradation by triple antibiotic paste and calcium hydroxide.

PURPOSE: To investigate and compare the effects of the two widely used regenerative endodontics medicaments: Triple antibiotic paste (ciprofloxacine-metronidazole-clindamycin) and calcium hydroxide on the microhardness and degradation of human root dentin. METHODS: Following ethical approval and subject consent to use teeth in this research study, 60 singled-rooted permanent human teeth were randomly divided into six groups:(1) Tri-antibiotic paste with distilled water, or with (2) propylene glycol, (3) calcium hydroxide with distilled water, (4) calcium hydroxide propylene glycol, (5) untreated extracted teeth as negative controls, or (6) teeth instrumented and filled with calcium hydroxide or tri-antibiotic paste as positive controls. The microhardness tests were conducted after 1 and 2 months of exposure to the medicaments using a Vickers microhardness tester. Raman spectroscopy and energy dispersive x-ray spectroscopy were used to evaluate the chemistry and structure of the root dentin. RESULTS: There were differences in the dentin microhardness following treatment with the medicaments or controls (P< 0.05). The time of root dentin exposure to the medicaments was similar (P> 0.05). The root dentin microhardness was lower in the teeth treated with the triple antibiotic paste or calcium hydroxide when combined with propylene glycol. The root dentin collagen in these treated teeth were also significantly degraded when viewed with Raman spectroscopy and energy dispersive x-ray spectroscopy, whereas the inorganic phase (dentin) remained unaltered. Samples exposed to the antimicrobial agents with water as a vehicle exhibited stronger microhardness and less degradation. CLINICAL SIGNIFICANCE: These ex vivo results suggest that the triple antibiotic paste and calcium hydroxide should be used with propylene glycol if a fast diffusion is desired or with water to avoid degrading the collagen and weakening the microhardness of the teeth. Clinical trials are needed of new formulations of medicaments with propylene glycol to disinfect teeth for regenerative endodontic procedures, to help strengthen the teeth to prevent the loss of children's permanent immature teeth by fracture following caries or trauma.

α-cyanobacteria possessing form IA RuBisCO globally dominate aquatic habitats.

RuBisCO (ribulose 1,5-bisphosphate carboxylase/oxygenase) is one the most abundant enzymes on Earth. Virtually all food webs depend on its activity to supply fixed carbon. In aerobic environments, RuBisCO struggles to distinguish efficiently between CO2 and O2. To compensate, organisms have evolved convergent solutions to concentrate CO2 around the active site. The genetic engineering of such inorganic carbon concentrating mechanisms (CCMs) into plants could help facilitate future global food security for humankind. In bacteria, the carboxysome represents one such CCM component, of which two independent forms exist: α and ß. Cyanobacteria are important players in the planet's carbon cycle and the vast majority of the phylum possess a ß-carboxysome, including most cyanobacteria used as laboratory models. The exceptions are the exclusively marine Prochlorococcus and Synechococcus that numerically dominate open ocean systems. However, the reason why marine systems favor an α-form is currently unknown. Here, we report the genomes of 58 cyanobacteria, closely related to marine Synechococcus that were isolated from freshwater lakes across the globe. We find all these isolates possess α-carboxysomes accompanied by a form 1A RuBisCO. Moreover, we demonstrate α-cyanobacteria dominate freshwater lakes worldwide. Hence, the paradigm of a separation in carboxysome type across the salinity divide does not hold true, and instead the α-form dominates all aquatic systems. We thus question the relevance of ß-cyanobacteria as models for aquatic systems at large and pose a hypothesis for the reason for the success of the α-form in nature.

Ab Initio Partition Functions and Thermodynamic Quantities for the Molecular Hydrogen Isotopologues.

In this work, we calculate the partition functions and thermodynamic quantities of molecular hydrogen isotopologues using the rovibrational energy levels provided by the highly accurate ab initio adiabatic potential energy functions recently determined by Pachucki and Komasa (Pachucki, K.; Komasa, J. J. Chem. Phys. 2014, 141, 224103). The partition functions are calculated by including all bound energy levels of the isotopologues, up to their dissociation limits, plus the quasi-bound levels lying below the centrifugal potential barriers. For the homonuclear isotopologues, H2, D2, and T2, we also determine the partition functions and thermodynamic quantities of the normal mixtures using the statistical treatment recently proposed by Colonna et al. (Colonna, G.; D'Angola, A.; Capitelli, M. Int. J. Hydrogen Energy 2012, 37, 9656) based on the definition of the partition function of the mixture, which avoids inconsistencies in the values of the thermodynamic quantities depending directly on the internal partition function, in the high-temperature limit.

On the Role of Entropy in the Stabilization of α-Helices.

Protein folding evolves by exploring the conformational space with a subtle balance between enthalpy and entropy changes which eventually leads to a decrease of free energy upon reaching the folded structure. A complete understanding of this process requires, therefore, a deep insight into both contributions to free energy. In this work, we clarify the role of entropy in favoring the stabilization of folded structures in polyalanine peptides with up to 12 residues. We use a novel method referred to as K2V that allows us to obtain the potential-energy landscapes in terms of residue conformations extracted from molecular dynamics simulations at conformational equilibrium and yields folding thermodynamic magnitudes, which are in agreement with the experimental data available. Our results demonstrate that the folded structures of the larger polyalanine chains are stabilized with respect to the folded structures of the shorter chains by both an energetic contribution coming from the formation of the intramolecular hydrogen bonds and an entropic contribution coming from an increase of the entropy of the solvent with approximate weights of 60 and 40%, respectively, thus unveiling a key piece in the puzzle of protein folding. In addition, the ability of the K2V method to provide the enthalpic and entropic contributions for individual residues along the peptide chain makes it clear that the energetic and entropic stabilizations are basically governed by the nearest neighbor residue conformations, with the folding propensity being rationalized in terms of triads of residues.

Intraresidual Correlated Motions in Peptide Chains.

We investigate the interresidual and intraresidual correlations between dihedral displacements of adjacent residues within model polyalanine peptides by analyzing extensive molecular dynamics trajectories. Correlations are evaluated individually at different residue conformations covering the whole (ϕi,ψi)-space. From these, we draw maps that unveil an unprecedented strong intramolecular correlation displaying opposite (correlated/anticorrelated) behaviors at different conformations. Both interresidual and intraresidual correlations arise from the propensity of the peptide to minimize the overall atomic displacements.

Tuning the Optical Properties of Novel Antitumoral Drugs Based on Cyclometalated Iridium(III) Complexes.

Most of the current efforts in drug discovery are devoted to the design of molecules able to mitigate side effects by concentrating the biological action in the targeted tissue. One promising strategy is photodynamic therapy, which is based on the in situ generation of reactive singlet oxygen upon radiation exposure. However, such an approach requires the use of an efficient photosensitizer. This contribution deals with the optical properties of an Ir(III) complex, [Ir(pbz)2(N^N)] (pbz = 2-phenylbenzimidazole; N^N = methyl 1-butyl-2-pyridyl-benzimidazole-5-carboxylate), which has recently been shown to exhort a strong photoactivity, but still needs further improvements to reach clinical applications. We performed density functional theory calculations at the M06, PBE0, ωB97xD, and CAM-B3LYP levels to predict the impact of introducing electron donor-acceptor groups into the nature of the lowest excited states. The simulations performed demonstrate that the presence of a NH2 at the pbz ligand and a NO2 group at the N^N ligand yield a bathochromic shift of absorption spectrum. We report the most sensitive positions to tune the optical signatures of this family of complexes.

Energetic Self-Folding Mechanism in α-Helices.

A novel energetic route driving the folding of a polyalanine peptide from an extended conformation to its α-helix native conformation is described, supported by a new method to compute mean potential energy surfaces accurately in terms of the dihedral angles of the peptide chain from extensive molecular dynamics simulations. The energetic self-folding (ESF) route arises specifically from the balance between the intrinsic propensity of alanine residues toward the αR conformation and two, opposite, effects: the destabilizing interaction with neighbor residues and the stabilizing formation of native hydrogen bonds, with the latter being dominant for large peptide lengths. The ESF mechanism provides simple but robust support to the nucleation-elongation or zipper models and offers a quantitative energetic funnel picture of the folding process. The mechanism is validated by the reasonable agreement between the computed folding energies and the experimental values.

Antibodies as Carrier Molecules: Encapsulating Anti-Inflammatory Drugs inside Herceptine.

The human epidermal growth factor receptor 2 (HER2) is overexpressed in about a third of breast cancer patients, with a strong involvement of the cyclooxygenase-2 (COX-2) enzyme in the tumor progress. HER2 and COX-2 are consequently potential targets for inhibiting carcionogenesis. Herceptin (trastuzumab) is an antibody that partially blocks HER2-positive cancers at their initial stage. Unfortunately, the overall response rate to the single treatment with this antibody is still modest, and therefore, it needs to be improved by combining several chemotherapeutic agents. On the other hand, nonsteroidal anti-inflammatory drugs (NSAIDs) are designed to halt COX-2 functionality, so they might also exhort an anticancer activity. In this contribution, dual Herceptin-NSAID drugs are designed using theoretical tools. More specifically, blind docking, molecular dynamics, and quantum calculations are performed to assess the stability of 14 NSAIDs embedded inside Herceptin. Our calculations reveal the feasibility of improving the antitumor activity of the parent Herceptin by designing a dual HER2-targeting with Etofenamate. That coupling mode might be used to further rationalize new clinical strategies beyond classical antibody dosages.

DFT Simulation of Structural and Optical Properties of 9-Aminoacridine Half-Sandwich Ru(II), Rh(III), and Ir(III) Antitumoral Complexes and Their Interaction with DNA.

In this work, we use DFT-based methods to simulate the chemical structures, optical properties, and interaction with DNA of a recently synthesized chelated C^N 9-aminoacridine arene Ru(II) anticancer agent and two new closely related Rh(III) and Ir(III) complexes using DFT-based methods. Four chemical models and a number of theoretical approaches, which representatively include the PBE0, B97D, ωB97X, ωB97X-D, M06, and M06-L density functionals and the LANL2DZ, def2-SVP, and def2-TZVP basis sets, are tested. The best overall accuracy/cost performance for the optimization process is reached at the ωB97X-D/def2-SVP and M06/def2-SVP levels of theory. Inclusion of explicit solvent molecules (CHCl3) further refines the geometry, while taking into account the crystal network gives no significant improvements of the computed bond distances and angles. The analysis of the excited states reveals that the M06 level matches better the experimental absorption spectra, compared to ωB97X-D. The use of the M06/def2-SVP approach is therefore a well-balanced method to study theoretically the bioactivity of this type of antitumoral complexes, so we couple this TD-DFT approach to molecular dynamics simulations in order to assess their reactivity with DNA. The reported results demonstrate that these drugs could be used to inject electrons into DNA, which might broaden their applications in photoactivated chemotherapy and as new materials for DNA-based electrochemical nanodevices.

Structure, Spectra, and DFT Simulation of Nickel Benzazolate Complexes with Tris(2-aminoethyl)amine Ligand.

Benzazolate complexes of Ni(II), [Ni(pbz)(tren)]ClO4 (pbz = 2-(2'-hydroxyphenyl)-benzimidazole (pbm), 1, 2-(2'-hydroxyphenyl)-benzoxazole (pbx), 2, 2-(2'-hydroxyphenyl)-benzothiazole (pbt), 3; tren = tris(2-aminoethyl)amine), are prepared by self-assembly reaction and structurally characterized. Theoretical DFT simulations are carried out to reproduce the features of their crystal structures and their spectroscopic and photophysic properties. The three complexes are moderately luminescent at room temperature both in acetonitrile solution and in the solid state. The simulations indicate that the absorption spectrum is dominated by two well-defined transitions, and the electronic density concentrates in three MOs around the benzazole ligands. The Stokes shifts of the emission spectra of complexes 1-3 are determined by optimizing the electronic excited state.

Understanding the connection between conformational changes of peptides and equilibrium thermal fluctuations.

Despite the increasing evidence that conformational transitions in peptides and proteins are driven by specific vibrational energy pathways along the molecule, the current experimental techniques of analysis do as yet not allow to study these biophysical processes in terms of anisotropic energy flows. Computational methods offer a complementary approach to obtain a more detailed understanding of the vibrational and conformational dynamics of these systems. Accordingly, in this work we investigate jointly the vibrational energy distribution and the conformational dynamics of trialanine peptide in water solution at room temperature by applying the Instantaneous Normal Mode analysis to the results derived from equilibrium molecular dynamics simulations. It is shown that conformational changes in trialanine are triggered by the vibrational energy accumulated in the low-frequency modes of the molecule, and that excitation is caused exclusively by thermal fluctuations of the solute-solvent system, thus excluding the possibility of an intramolecular vibrational energy redistribution process.

Relaxation pathways of the OD stretch fundamental of HOD in liquid H2O.

The molecular dynamics with quantum transitions method is used to study the vibrational relaxation of the OD stretching mode of HOD dissolved in liquid H2O water at 303 K. All the vibrational modes of the solute and solvent molecules that participate in the relaxation process are described by quantum mechanics, while the rotational and translational degrees of freedom are treated classically. A modification of the water intramolecular SPC/E (Simple Point Charge/Extended) force field providing vibrational frequencies in solution closer to the experimental values is proposed to analyze the influence of the vibrational energy gaps on the relaxation channels. The relaxation times obtained are in satisfactory agreement with experimental values. The energy transfer during the relaxation process alters significantly the H-bond network around the HOD molecule. The analysis of the vibrational transitions during the relaxation process reveals a complex mechanism which involves the participation of both intra- and intermolecular channels and provides a compromise for the different interpretations of the experimental data reported for this system in recent years.

Conformational Changes of Trialanine in Water Induced by Vibrational Relaxation of the Amide I Mode.

Most of the protein-based diseases are caused by anomalies in the functionality and stability of these molecules. Experimental and theoretical studies of the conformational dynamics of proteins are becoming in this respect essential to understand the origin of these anomalies. However, a description of the conformational dynamics of proteins based on mechano-energetic principles still remains elusive because of the intrinsic high flexibility of the peptide chains, the participation of weak noncovalent interactions, and the role of the ubiquitous water solvent. In this work, the conformational dynamics of trialanine dissolved in water (D2O) is investigated through Molecular Dynamics (MD) simulations combined with instantaneous normal modes (INMs) analysis both at equilibrium and after the vibrational excitation of the C-terminal amide I mode. The conformational equilibrium between α and pPII conformers is found to be altered by the intramolecular relaxation of the amide I mode as a consequence of the different relaxation pathways of each conformer which modify the amount of vibrational energy stored in the torsional motions of the tripeptide, so the α → pPII and pPII → α conversion rates are increased differently. The selectivity of the process comes from the shifts of the vibrational frequencies with the conformational changes that modify the resonance conditions driving the intramolecular energy flows.

Mutagenic effects induced by the attack of NO2 radical to the guanine-cytosine base pair.

We investigate the attack of the nitrogen dioxide radical (NO(•) 2) to the guanine-cytosine (GC) base pair and the subsequent tautomeric reactions able to induce mutations, by means of density functional theory (DFT) calculations. The conducted simulations allow us to identify the most reactive sites of the GC base pair. Indeed, the computed relative energies demonstrate that the addition of the NO(•) 2 radical to the C8 position of the guanine base forms to the most stable adduct. Although the initial adducts might evolve to non-canonical structures via inter-base hydrogen bonds rearrangements, the probability for the proton exchange to occur lies in the same range as that observed for undamaged DNA. As a result, tautomeric errors in NO2-attacked DNA arises at the same rate as in canonical DNA, with no macroscopic impact on the overall stability of DNA. The potential mutagenic effects of the GC-NO(•) 2 radical adducts likely involve side reactions, e.g., the GC deprotonation to the solvent, rather than proton exchange between guanine and cytosine basis.