An ortholog of the Vasa intronic gene is required for small RNA-mediated translation repression in .

Small RNAs (sRNAs) associate with Argonaute (AGO) proteins in effector complexes, termed RNA-induced silencing complexes (RISCs), which regulate complementary transcripts by translation inhibition and/or RNA degradation. In the unicellular alga , several metazoans, and land plants, emerging evidence indicates that polyribosome-associated transcripts can be translationally repressed by RISCs without substantial messenger RNA (mRNA) destabilization. However, the mechanism of translation inhibition in a polyribosomal context is not understood.

The NMR solution structure and function of RPA3313: a putative ribosomal transport protein from Rhodopseudomonas palustris.

Protein function elucidation often relies heavily on amino acid sequence analysis and other bioinformatics approaches. The reliance is extended to structure homology modeling for ligand docking and protein-protein interaction mapping. However, sequence analysis of RPA3313 exposes a large, unannotated class of hypothetical proteins mostly from the Rhizobiales order.

Glucose Metabolism and AMPK Signaling Regulate Dopaminergic Cell Death Induced by Gene (α-Synuclein)-Environment (Paraquat) Interactions.

While environmental exposures are not the single cause of Parkinson's disease (PD), their interaction with genetic alterations is thought to contribute to neuronal dopaminergic degeneration. However, the mechanisms involved in dopaminergic cell death induced by gene-environment interactions remain unclear. In this work, we have revealed for the first time the role of central carbon metabolism and metabolic dysfunction in dopaminergic cell death induced by the paraquat (PQ)-α-synuclein interaction.

Transient sampling of aggregation-prone conformations causes pathogenic instability of a parkinsonian mutant of DJ-1 at physiological temperature.

Various missense mutations in the cytoprotective protein DJ-1 cause rare forms of inherited parkinsonism. One mutation, M26I, diminishes DJ-1 protein levels in the cell but does not result in large changes in the three-dimensional structure or thermal stability of the protein. Therefore, the molecular defect that results in loss of M26I DJ-1 protective function is unclear.

Comparison of Mannose, Ethylene Glycol, and Methoxy-Terminated Diluents on Specificity and Selectivity of Electrochemical Peptide-Based Sensors.

We report the synthesis and application of three new antifouling diluents for the fabrication of an E-PB HIV sensor. Among the three thiolated antifouling diluents used in this study, the methoxy-terminated diluent (C6-MEG) is the most effective in alleviating both nonspecific binding and adsorption of matrix contaminants onto the sensor surface, especially when compared to the mannose- (C6-MAN) and ethylene-glycol-terminated (C6-EG) diluents. The sensor fabricated with C6-MEG has a specificity factor (∼13.

Sphingolipid metabolism is strikingly different between pollen and leaf in Arabidopsis as revealed by compositional and gene expression profiling.

Although sphingolipids are essential for male gametophytic development in Arabidopsis thaliana, sphingolipid composition and biosynthetic gene expression have not been previously examined in pollen. In this report, electrospray ionization (ESI)-MS/MS was applied to characterization of sphingolipid compositional profiles in pollen isolated from wild type Arabidopsis Col-0 and a long-chain base (LCB) Δ4 desaturase mutant. Pollen fractions were highly enriched in glucosylceramides (GlcCer) relative to levels previously reported in leaves.

Iterative assembly of two separate polyketide chains by the same single-module bacterial polyketide synthase in the biosynthesis of HSAF.

Antifungal HSAF (heat-stable antifungal factor, dihydromaltophilin) is a polycyclic tetramate macrolactam from the biocontrol agent Lysobacter enzymogenes. Its biosynthetic gene cluster contains only a single-module polyketide synthase-nonribosomal peptide synthetase (PKS-NRPS), although two separate hexaketide chains are required to assemble the skeleton. To address the unusual biosynthetic mechanism, we expressed the biosynthetic genes in two "clean" strains of Streptomyces and showed the production of HSAF analogues and a polyene tetramate intermediate.

Tricarballylic ester formation during biosynthesis of fumonisin mycotoxins in .

Fumonisins are agriculturally important mycotoxins produced by the maize pathogen The chemical structure of fumonisins contains two tricarballylic esters, which are rare structural moieties and important for toxicity. The mechanism for the tricarballylic ester formation is not well understood. gene of was predicted to encode a dehydrogenase/reductase, and when it was deleted, the mutant produced tetradehydro fumonisins (DH-FB).

Overexpression of human fatty acid transport protein 2/very long chain acyl-CoA synthetase 1 (FATP2/Acsvl1) reveals distinct patterns of trafficking of exogenous fatty acids.

In mammals, the fatty acid transport proteins (FATP1 through FATP6) are members of a highly conserved family of proteins, which function in fatty acid transport proceeding through vectorial acylation and in the activation of very long chain fatty acids, branched chain fatty acids and secondary bile acids. FATP1, 2 and 4, for example directly function in fatty acid transport and very long chain fatty acids activation while FATP5 does not function in fatty acid transport but activates secondary bile acids. In the present work, we have used stable isotopically labeled fatty acids differing in carbon length and saturation in cells expressing FATP2 to gain further insights into how this protein functions in fatty acid transport and intracellular fatty acid trafficking.

Development of affinity microcolumns for drug-protein binding studies in personalized medicine: interactions of sulfonylurea drugs with in vivo glycated human serum albumin.

This report used high-performance affinity microcolumns to examine the changes in binding by sulfonylurea drugs to in vivo glycated HSA that had been isolated from individual patients with diabetes. An immunoextraction approach was developed to isolate HSA and glycated HSA from clinical samples, using only 20 μL of plasma or serum and 6-12 nmol of protein to prepare each affinity microcolumn. It was found that the affinity microcolumns could be used in either frontal analysis or zonal elution studies, which typically required only 4-8 min per run.

Knockdown of the cellular protein LRPPRC attenuates HIV-1 infection.

HIV-1 exploits numerous host cellular pathways for productive infection. To identify novel factors involved in HIV-1 replication, HIV-1 integrase and matrix protein complexes were captured at 4 hours post infection for proteomic analysis using an affinity purification system. Leucine-rich PPR-motif containing (LRPPRC) protein, a cellular protein involved in mitochondrial function, cell metabolism, and cell-cycle progression was identified as one of the candidate HIV-1 factors.

The applicability of molecular descriptors for designing an electrospray ionization mass spectrometry compatible library for drug discovery.

Detecting a small molecular-weight compound by electrospray ionization mass spectrometry (ESI-MS) requires the compound to obtain a charge. Factors such as gas-phase proton affinities and analyte surface activity are correlated with a positive ESI-MS response, but unfortunately it is extremely challenging to predict from a chemical structure alone if a compound is likely to yield an observable molecular-ion peak in an ESI-MS spectrum. Thus, the design of a chemical library for an ESI-MS ligand-affinity screen is particularly daunting.

Paramecium bursaria chlorella virus 1 proteome reveals novel architectural and regulatory features of a giant virus.

The 331-kbp chlorovirus Paramecium bursaria chlorella virus 1 (PBCV-1) genome was resequenced and annotated to correct errors in the original 15-year-old sequence; 40 codons was considered the minimum protein size of an open reading frame. PBCV-1 has 416 predicted protein-encoding sequences and 11 tRNAs. A proteome analysis was also conducted on highly purified PBCV-1 virions using two mass spectrometry-based protocols.

Unusual activities of the thioesterase domain for the biosynthesis of the polycyclic tetramate macrolactam HSAF in Lysobacter enzymogenes C3.

HSAF is an antifungal natural product with a new mode of action. A rare bacterial iterative PKS-NRPS assembles the HSAF skeleton. The biochemical characterization of the NRPS revealed that the thioesterase (TE) domain possesses the activities of both a protease and a peptide ligase.

Structure function analysis of an ADP-ribosyltransferase type III effector and its RNA-binding target in plant immunity.

The Pseudomonas syringae type III effector HopU1 is a mono-ADP-ribosyltransferase that is injected into plant cells by the type III protein secretion system. Inside the plant cell it suppresses immunity by modifying RNA-binding proteins including the glycine-rich RNA-binding protein GRP7. The crystal structure of HopU1 at 2.

Human fatty acid transport protein 2a/very long chain acyl-CoA synthetase 1 (FATP2a/Acsvl1) has a preference in mediating the channeling of exogenous n-3 fatty acids into phosphatidylinositol.

The trafficking of fatty acids across the membrane and into downstream metabolic pathways requires their activation to CoA thioesters. Members of the fatty acid transport protein/very long chain acyl-CoA synthetase (FATP/Acsvl) family are emerging as key players in the trafficking of exogenous fatty acids into the cell and in intracellular fatty acid homeostasis. We have expressed two naturally occurring splice variants of human FATP2 (Acsvl1) in yeast and 293T-REx cells and addressed their roles in fatty acid transport, activation, and intracellular trafficking.

Quantitative analysis of glycation patterns in human serum albumin using 16O/18O-labeling and MALDI-TOF MS.

Background: The glycation of human serum albumin (HSA) during diabetes can affect the ability of this protein to bind drugs and small solutes in blood. This study describes the use of (16)O/(18)O-labeling and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry to compare the levels of modification that occur throughout HSA under various glycation conditions in vitro. These quantitative studies build on a recent report that has identified the early and advanced glycation products that are formed on such samples of HSA.

A cytoplasm-specific activity encoded by the Trithorax-like ATX1 gene.

Eukaryotes produce multiple products from a single gene locus by alternative splicing, translation or promoter usage as mechanisms expanding the complexity of their proteome. Trithorax proteins, including the Arabidopsis Trithorax-like protein ATX1, are histone modifiers regulating gene activity. Here, we report that a novel member of the Trithorax family has a role unrelated to chromatin.

Fluoride-Promoted Ligand Exchange in Diaryliodonium Salts.

Diaryliodonium salts are shown to undergo rapid, fluoride-promoted aryl exchange reactions at room temperature in acetonitrile. Aryl exchange is shown to be exquisitely sensitive to the concentration of fluoride ion in solution; fast exchange is observed as the fluoride concentration approaches a stoichiometric amount at 50 mM substrate concentration. The reaction is slowed, but not halted if benzene is the solvent, indicating that free fluoride ion or a four-coordinate anionic I(III) species may be responsible for the exchange.

Comparison of modification sites formed on human serum albumin at various stages of glycation.

Background: Many of the complications encountered during diabetes can be linked to the non-enzymatic glycation of proteins, including human serum albumin (HSA). However, there is little information regarding how the glycation pattern of HSA changes as the total extent of glycation is varied. The goal of this study was to identify and conduct a semi-quantitative comparison of the glycation products on HSA that are produced in the presence of various levels of glycation.

Characterization of surface-exposed reactive cysteine residues in Saccharomyces cerevisiae.

Numerous cellular processes are subject to redox regulation, and thiol-dependent redox control, acting through reactive cysteine (Cys) residues, is among the major mechanisms of redox regulation. However, information on the sets of proteins that provide thiol-based redox regulation or are affected by it is limited. Here, we describe proteomic approaches to characterize proteins that contain reactive thiols and methods to identify redox Cys in these proteins.

Improved arene fluorination methodology for I(III) salts.

The use of low polarity aromatic solvents (benzene or toluene) and/or the removal of inorganic salts results in dramatically improved yields of fluorinated arenes from diaryliodonium salts. This methodology is shown to "scale down" to the conditions used typically for radiotracer synthesis.

Proteomic analysis of atrazine exposure in Drosophila melanogaster (Diptera: Drosophilidae).

Atrazine is a widely used herbicide that has been reported to induce the activity of certain detoxification enzymes and to affect insecticide toxicity in organisms experiencing simultaneous exposure to both atrazine and insecticides. In this study, the effects of atrazine (2-chloro-4-ethylamino-6-isopropylamino-1,3,5-triazine) exposure on protein expression in male and female Drosophila melanogaster adults in both microsomal and cytosolic cell fractions was investigated by 2-dimensional gel electrophoresis. Differentially expressed proteins (vs.