Light-Induced Nanosecond Relaxation Dynamics of Rhenium-Labeled Pseudomonas aeruginosa Azurins.

Time-resolved phosphorescence spectra of Re(CO)3(dmp)+ and Re(CO)3(phen)+ chromophores (dmp = 4,7-dimethyl-1,10-phenanthroline, phen = 1,10-phenanthroline) bound to surface histidines (H83, H124, and H126) of Pseudomonas aeruginosa azurin mutants exhibit dynamic band maxima shifts to lower wavenumbers following 3-exponential kinetics with 1-5 and 20-100 ns major phases and a 1.1-2.5 μs minor (5-16%) phase. Observation of slow relaxation components was made possible by using an organometallic Re chromophore as a probe whose long phosphorescence lifetime extends the observation window up to ∼3 μs. Integrated emission-band areas also decay with 2- or 3-exponential kinetics; the faster decay phase(s) is relaxation-related, whereas the slowest one [360-680 ns (dmp); 90-140 ns (phen)] arises mainly from population decay. As a result of shifting bands, the emission intensity decay kinetics depend on the detection wavelength. Detailed kinetics analyses and comparisons with band-shift dynamics are needed to disentangle relaxation and population decay kinetics if they occur on comparable timescales. The dynamic phosphorescence Stokes shift in Re-azurins is caused by relaxation motions of the solvent, the protein, and solvated amino acid side chains at the Re binding site in response to chromophore electronic excitation. Comparing relaxation and decay kinetics of Re(dmp)124K122Cu II and Re(dmp)124W122Cu II suggests that electron transfer (ET) and relaxation motions in the W122 mutant are coupled. It follows that nanosecond and faster photo-induced ET steps in azurins (and likely other redox proteins) occur from unrelaxed systems; importantly, these reactions can be driven (or hindered) by structural and solvational dynamics

Two-photon spectroscopy of tungsten(0) arylisocyanides using nanosecond-pulsed excitation

The two-photon absorption (TPA) cross sections (δ) for tungsten(0) arylisocyanides (W(CNAr)6) were determined in the 800–1000 nm region using two-photon luminescence (TPL) spectroscopy. The complexes have high TPA cross sections, in the range 1000–2000 GM at 811.8 nm. In comparison, the cross section at 811.8 nm for tris-(2,2′-bipyridine)ruthenium(II), [Ru(bpy)_3]^(2+), is 7 GM. All measurements were performed using a nanosecond-pulsed laser system

Hole Hopping Across a Protein-Protein Interface.

We have investigated photoinduced hole hopping in a Pseudomonas aeruginosa azurin mutant Re126WWCuI, where two adjacent tryptophan residues (W124 and W122) are inserted between the CuI center and a Re photosensitizer coordinated to a H126 imidazole (Re = ReI(H126)(CO)3(dmp)+, dmp = 4,7-dimethyl-1,10-phenanthroline). Optical excitation of this mutant in aqueous media (//(CuII)' back ET that occurs over 12 Å, in contrast to the 23 Å, 120 us step in Re126WWCuI. Importantly, dimerization makes Re126FWCuI photoreactive and, in the case of {Re126WWCuI}2, channels the photoproduced "hole" to the molecule that was not initially photoexcited, thereby shortening the lifetime of ReI(H126)(CO)3(dmp•-)//CuII. Whereas two adjacent W124 and W122 indoles dramatically enhance CuI->*Re intramolecular multistep ET, the tryptophan quadruplex in {Re126WWCuI}2 does not accelerate intermolecular electron transport; instead, it acts as a hole storage and crossover unit between inter- and intramolecular ET pathways. Irradiation of {Re126WWCuII}2 or {Re126FWCuII}2 also triggers intermolecular *Re////(W122•+)' intermolecular charge recombination. Our findings shed light on the factors that control interfacial hole/electron hopping in protein complexes and on the role of aromatic amino acids in accelerating long-range electron transport

Estudo da atividade inseticida de algumas espécies vegetais.

Foram estudadas 30 espécies vegetais, quanto a atividade inseticida. A partir das diferentes partes desses vegetais foram preparados 240 extratos, utilizando hexano, acetona, etanol 96 e 30 GL, como líquidos extratores. Os testes foram efetuados contra os insetos: Musca domestica, Ceratitis capitata Zabrotes subfasciatus, Spodoptera frugiperda e Anthonomus grandis. Dos vegetais testados, os melhores resultados foram apresentados pelos extratos: hexanico de sementes de Annona crassiflora, hidroalcoolico de raiz de potomorphe umbellata, contra Ceratilis capitata e zabrotes subfasciatus, respectivamente; extratos acetônicos de sementes de Annona cacans e A. squamosa contra Musca domestica; extrato hexanico de sementes Carpotroche brasiliensis contra Ceratilis capitata e extratos etanolicos de frutos e sementes de A. squamosa contra Spodoptera frugiperda. Entre os vegetais que apresentaram atividade, observou-se predominância de espécies da família Annonaceae

Two Tryptophans Are Better Than One in Accelerating Electron Flow through a Protein

We have constructed and structurally characterized a <i>Pseudomonas aeruginosa</i> azurin mutant <b>Re126WWCu^I</b>, where two adjacent tryptophan residues (W124 and W122, indole separation 3.6–4.1 Å) are inserted between the Cu^I center and a Re photosensitizer coordinated to the imidazole of H126 (Re^I(H126)­(CO)₃(4,7-dimethyl-1,10-phenanthroline)⁺). Cu^I oxidation by the photoexcited Re label (*Re) 22.9 Å away proceeds with a ∼70 ns time constant, similar to that of a single-tryptophan mutant (∼40 ns) with a 19.4 Å Re–Cu distance. Time-resolved spectroscopy (luminescence, visible and IR absorption) revealed two rapid reversible electron transfer steps, W124 → *Re (400–475 ps, <i>K</i>₁ ≅ 3.5–4) and W122 → W124^•+ (7–9 ns, <i>K</i>₂ ≅ 0.55–0.75), followed by a rate-determining (70–90 ns) Cu^I oxidation by W122^•+ ca. 11 Å away. The photocycle is completed by 120 μs recombination. No photochemical Cu^I oxidation was observed in <b>Re126FWCu^I</b>, whereas in <b>Re126WFCu^I</b>, the photocycle is restricted to the ReH126W124 unit and Cu^I remains isolated. QM/MM/MD simulations of <b>Re126WWCu^I</b> indicate that indole solvation changes through the hopping process and W124 → *Re electron transfer is accompanied by water fluctuations that tighten W124 solvation. Our finding that multistep tunneling (hopping) confers a ∼9000-fold advantage over single-step tunneling in the double-tryptophan protein supports the proposal that hole-hopping through tryptophan/tyrosine chains protects enzymes from oxidative damage

Downsizing a human inflammatory protein to a small molecule with equal potency and functionality

A significant challenge in chemistry is to rationally reproduce the functional potency of a protein in a small molecule, which is cheaper to manufacture, non-immunogenic, and also both stable and bioavailable. Synthetic peptides corresponding to small bioactive protein surfaces do not form stable structures in water and do not exhibit the functional potencies of proteins. Here we describe a novel approach to growing small molecules with protein-like potencies from a functionally important amino acid of a protein. A 77-residue human inflammatory protein (complement C3a) important in innate immunity is rationally transformed to equipotent small molecules, using peptide surrogates that incorporate a turn-inducing heterocycle with correctly positioned hydrogen-bond-accepting atoms. Small molecule agonists (molecular weigh

Correlation Index-Based Responsible-Enzyme Gene Screening (CIRES), a Novel DNA Microarray-Based Method for Enzyme Gene Involved in Glycan Biosynthesis

BACKGROUND: Glycan biosynthesis occurs though a multi-step process that requires a variety of enzymes ranging from glycosyltransferases to those involved in cytosolic sugar metabolism. In many cases, glycan biosynthesis follows a glycan-specific, linear pathway. As glycosyltransferases are generally regulated at the level of transcription, assessing the overall transcriptional profile for glycan biosynthesis genes seems warranted. However, a systematic approach for assessing the correlation between glycan expression and glycan-related gene expression has not been reported previously. METHODOLOGY: To facilitate genetic analysis of glycan biosynthesis, we sought to correlate the expression of genes involved in cell-surface glycan formation with the expression of the glycans, as detected by glycan-recognizing probes. We performed cross-sample comparisons of gene expression profiles using a newly developed, glycan-focused cDNA microarray. Cell-surface glycan expression profiles were obtained using flow cytometry of cells stained with plant lectins. Pearson's correlation coefficients were calculated for these profiles and were used to identify enzyme genes correlated with glycan biosynthesis. CONCLUSIONS: This method, designated correlation index-based responsible-enzyme gene screening (CIRES), successfully identified genes already known to be involved in the biosynthesis of certain glycans. Our evaluation of CIRES indicates that it is useful for identifying genes involved in the biosynthesis of glycan chains that can be probed with lectins using flow cytometry

Direct detection of the nitrooxyperoxy radicals in the oxidation of VOCs by NO_3

The photooxidn. of volatile org. compds. (VOCs) is a significant source of secondary org. aerosols (SOA) in the troposphere. The nighttime chem. leading to SOA growth however remains less characterized. The nitrate radical NO3 is one of the most important nighttime oxidants. In the presence of oxygen, the free radical addn. of NO3 to unsatd. VOCs will lead to formation of nitrooxyalkyl peroxy radicals. While various studies have measured the further product yields and SOA formation from the reaction, the peroxy intermediate has yet to be directly obsd. We use cavity ringdown spectroscopy to directly detect the nitrooxyalkyl peroxy radicals via the A-X electronic absorption. The A-X electronic transition has enough structure to distinguish between different peroxy radicals as well as different conformers of the same peroxy radical. In order to elucidate the mechanism and formation of the intermediate, we present results from the addn. of NO3 to one of the simplest unsatd. VOCs, cis and trans 2-butene