Codon usage in the vertebrate hemoglobins and its implications

A study of codon usage in vertebrate hemoglobins revealed an evolutionary trend toward elevated numbers of CpG codon boundary pairs in mammalian hemoglobin alpha genes. Selection for CpG codon boundaries countering the generally observed CpG suppression is strongly suggested by these data. These observations parallel recently published experimental results that indicate that constitutive expression of the human alpha-globin gene appears to be determined by regulatory information encoded within the structural gene. The possibility is raised that, in the absence of selection, CpG decay can be used to date the evolutionary origin of a mammalian alpha pseudogene from its active alpha gene.      

Soil moisture, no-tillage and predator effects on southern corn rootworm survival in peanut agroecosystems

Experiments examined the effects of no-tillage (NT) and conventional-tillage (CT) and increased soil moisture (irrigation) on southern corn rootworm (SCR), Diabrotica undecimpunctata howardi Barber (Coleoptera: Chrysomelidae), oviposition and survival and SCR below-ground predators in peanut (Arachis hypogeae L.) agroecosystems. No-tillage systems without irrigation had significantly greater soil moisture than CT systems without irrigation in 1987, but not in 1988. Both moist soil and the presence of an organic residue increased the number of southern corn rootworm eggs in a system. The same 17 species of predatory arthropods were found in all systems. However, their activity in the top 10-cm of soil which increased in NT systems, determined how effective they were as predators. Only 7 predators significantly reduced SCR immature stages, and of these 7, only one, Formicidae, preyed on all stages of SCR. Southern corn rootworm first instars and eggs had the greatest overall percent mortality, with second and third instars the lowest. Peanut yields and pod quality were equal between NT and CT systems. Overall return per ha showed NT, no-irrigation systems with the greatest return. Abiotic parameters dominated SCR survival in CT systems, while biotic parameters had the greatest influence on SCR survival in NT systems.

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Résumé Des expériences ont été entreprises pour examiner l'influence de labours classiques ou de leur absence et de l'augmentation de l'humidité du sol (irrigation) sur la ponte et la survie de D. undecimpunctata howardi Barber (Coleo. Chrysomelidae) et sur ses prédateurs souterrains dans des agrosystèmes à arachide. L'humidité du sol s'est montrée significativement plus élevée dans les parcelles irriguées pendant les deux années, 1987 et 1988, mais, en 1987 et non en 1988, l'humidité du sol a été significativement plus élevée dans le sol non labouré pourtant non irrigué que dans le sol labouré et irrigué. La présence de sol humide et de résidus organiques augmente le nombre d'ufs émis. Les mêmes 17 espèces d'arthropodes prédateurs dateurs souterrains ont été rencontrés dans toutes les sols non labourés et parfois dans les sols labourés irrigués; leur activité détermine leur efficacité comme prédateur. 7 espèces réduisent les effectifs de larves, mais une seule d'entre elles, une Formicidae, (Lasius spp. et Pheidole spp.) s'est révélée prédatrice de tous les stades de la chrysomèle. La plus forte mortalité de D. u. howardi a été observée sur les ufs et les larves de premier stade, la plus faible sur les larves de deuxième et troisième stade. La récolte et la qualité des arachides sont identiques avec ou sans labour. Ainsi, la rentabilité financière par ha était supérieure dans les systèmes sans labour non irrigués. Il découle de cette étude que les paramètres physiques dominent la survie de la chrysomèle dans les systèmes avec labour, tandis que les paramètres biologiques influent le plus sur la survie dans les systèmes sans labour.

     

Synthesis and biological evaluation of 18F labeled fluoro-oligo-ethoxylated 4-benzylpiperazine derivatives for sigma-1 receptor imaging

We report the synthesis and evaluation of a series of fluoro-oligo-ethoxylated 4-benzylpiperazine derivatives as potential σ₁ receptor ligands. In vitro competition binding assays showed that 1-(1,3-benzodioxol-5-ylmethyl)-4-(4-(2-fluoroethoxy)benzyl)piperazine (6) exhibits low nanomolar affinity for σ₁ receptors (K_i = 1.85 ± 1.59 nM) and high subtype selectivity (σ₂ receptor: K_i = 291 ± 111 nM; K_iσ₂/K_iσ₁ = 157). [¹⁸F]6 was prepared in 30–50% isolated radiochemical yield, with radiochemical purity of >99% by HPLC analysis after purification, via nucleophilic ¹⁸F^? substitution of the corresponding tosylate precursor. The log D_{pH 7.4} value of [¹⁸F]6 was found to be 2.57 ± 0.10, which is within the range expected to give high brain uptake. Biodistribution studies in mice demonstrated relatively high concentration of radiotracers in organs known to contain σ₁ receptors, including the brain, lungs, kidneys, heart, and spleen. Administration of haloperidol 5 min prior to injection of [¹⁸F]6 significantly reduced the concentration of radiotracers in the above-mentioned organs. The accumulation of radiotracers in the bone was quite low suggesting that [¹⁸F]6 is relatively stable to in vivo defluorination. The ex vivo autoradiography in rat brain showed high accumulation of radiotracers in the brain areas known to possess high expression of σ₁ receptors. These findings suggest that [¹⁸F]6 is a suitable radiotracer for imaging σ₁ receptors with PET in vivo.     

How do dilatations form in mosquito ovarioles?

Ovariolar dilatations have been used to estimate parity status in mosquitoes for over 40 years. The theoretical basis for this parity-diagnostic method, which was established for mosquitoes by Polovodova 1, has been challenged by a number of Russian researchers. The findings of this new school of researchers have elicited little response from those currently using the method outside Russia, except for a recent paper by Hoc and Charlwood2. This has stimulated Andrew Fox and Reinhart Brust to review the formation of ovariolar dilatations here, and compare what they term the Old and the New Schools. Although the basic application of Polovodova's method is unaffected by the premise of either School, a clear understanding of ovarian dynamics is needed to understand the method and its limits.     

Flavobacterium johnsoniae GldH is a lipoprotein that is required for gliding motility and chitin utilization

Cells of Flavobacterium johnsoniae move rapidly over surfaces by gliding motility. The mechanism of this form of motility is not known. Six genes (gldA, gldB, gldD, gldF, gldG, and ftsX) that are required for gliding have been described. Tn4351 mutagenesis was used to identify another gene, gldH, which is required for cell movement. GldH mutants formed nonspreading colonies, and individual cells lacked the cell movements and ability to propel latex spheres along their surfaces that are characteristic of wild-type cells. gldH mutants also failed to digest chitin and were resistant to bacteriophages that infect wild-type cells. Introduction of pMM293, which carries wild-type gldH, restored to the gldH mutants colony spreading, cell motility, the ability to move latex spheres, phage sensitivity, and the ability to digest chitin. gldH encodes a predicted 141-amino-acid protein that localized to the membrane fraction. Labeling studies with [3H]palmitate demonstrated that GldH is a lipoprotein. GldB and GldD, which were previously described, also appear to be lipoproteins. GldH does not exhibit significant amino acid similarity to proteins of known function in the databases. Putative homologs of gldH of unknown function are found in motile (Cytophaga hutchinsonii) and apparently nonmotile (Bacteroides thetaiotaomicron, Bacteroides fragilis, Tannerella forsythensis, Porphyromonas gingivalis, and Prevotella intermedia) members of the Cytophaga-Flavobacterium-Bacteroides group.     

Mycobacterium tuberculosis lipolytic enzymes as potential biomarkers for the diagnosis of active tuberculosis

Background

New diagnosis tests are urgently needed to address the global tuberculosis (TB) burden and to improve control programs especially in resource-limited settings. An effective in vitro diagnostic of TB based on serological methods would be regarded as an attractive progress because immunoassays are simple, rapid, inexpensive, and may offer the possibility to detect cases missed by standard sputum smear microscopy. However, currently available serology tests for TB are highly variable in sensitivity and specificity. Lipolytic enzymes have recently emerged as key factors in lipid metabolization during dormancy and/or exit of the non-replicating growth phase, a prerequisite step of TB reactivation. The focus of this study was to analyze and compare the potential of four Mycobacterium tuberculosis lipolytic enzymes (LipY, Rv0183, Rv1984c and Rv3452) as new markers in the serodiagnosis of active TB.

Methods

Recombinant proteins were produced and used in optimized ELISA aimed to detect IgG and IgM serum antibodies against the four lipolytic enzymes. The capacity of the assays to identify infection was evaluated in patients with either active TB or latent TB and compared with two distinct control groups consisting of BCG-vaccinated blood donors and hospitalized non-TB individuals.

Results

A robust humoral response was detected in patients with active TB whereas antibodies against lipolytic enzymes were infrequently detected in either uninfected groups or in subjects with latent infection. High specifity levels, ranging from 93.9% to 97.5%, were obtained for all four antigens with sensitivity values ranging from 73.4% to 90.5%, with Rv3452 displaying the highest performances. Patients with active TB usually exhibited strong IgG responses but poor IgM responses.

Conclusion

These results clearly indicate that the lipolytic enzymes tested are strongly immunogenic allowing to distinguish active from latent TB infections. They appear as potent biomarkers providing high sensitivity and specificity levels for the immunodiagnosis of active TB.     

The peptide route to multifunctional gold nanoparticles

Extremely stable, peptide-capped gold nanoparticles with two different biomolecular recognition motifs expressed on their surface have been prepared, and their specific and selective binding to artificial, DNA-modified target particles and to DNA and protein microarrays has been demonstrated. Stabilization and biofunctionalization has been achieved in a single preparative step starting with citrate-stabilized gold hydrosols and a derivatization cocktail of peptide-capping ligands, which carry the functionalities of choice.     

3D QSAR study, synthesis, and in vitro evaluation of (+)-5-FBVM as potential PET radioligand for the vesicular acetylcholine transporter (VAChT)

Located in presynaptic cholinergic nerve terminals, the vesicular acetylcholine transporter (VAChT) represents a potential target for quantitative visualization of early degeneration of cholinergic neurons in Alzheimer's disease using PET. Benzovesamicol derivatives are proposed as radioligands for this purpose. We report QSAR studies of vesamicol and benzovesamicol derivatives taking into account the stereoselectivity of the VAChT binding site. Use of different data sets and different models in this study revealed that both enantiomers of 5-fluoro-3-(4-phenyl-piperidin-1-yl)-1,2,3,4-tetrahydro-naphthalen-2-ol (5-FBVM) are promising candidates, with predicted VAChT affinities between 6.1 and 0.05 nM. The synthesis of enantiopure (R,R)- and (S,S)-5-FBVM and their corresponding triazene precursors for future radiofluorination is reported. Both enantiomers exhibited high in vitro affinity for VAChT [(+)-5-FBVM: K(i)=6.95 nM and (-)-5-FBVM: K(i)=3.68 nM] and were selective for σ(2) receptors (～70-fold), only (+)-5-FBVM is selective for σ(1) receptors (～fivefold). These initial results suggest that (+)-(S,S)-5-FBVM warrants further investigation as a potential radioligand for in vivo PET imaging of cholinergic nerve terminals.     

Exposure of mycobacteria to cell wall-inhibitory drugs decreases production of arabinoglycerolipid related to Mycolyl-arabinogalactan-peptidoglycan metabolism

The "cell wall core" consisting of a mycolyl-arabinogalactan-peptidoglycan (mAGP) complex represents the hallmark of the mycobacterial cell envelope. It has been the focus of intense research at both structural and biosynthetic levels during the past few decades. Because it is essential, mAGP is also regarded as a target for several antitubercular drugs. Herein, we demonstrate that exposure of Mycobacterium bovis Bacille Calmette-Guérin or Mycobacterium marinum to thiacetazone, a second line antitubercular drug, is associated with a severe decrease in the level of a major apolar glycolipid. This inhibition requires MmaA4, a methyltransferase reported to participate in the activation process of thiacetazone. Following purification, this glycolipid was subjected to detailed structural analyses, combining gas-liquid chromatography, mass spectrometry, and nuclear magnetic resonance. This allowed to identify it as a 5-O-mycolyl-β-Araf-(1→2)-5-O-mycolyl-α-Araf-(1→1)-Gro, designated dimycolyl diarabinoglycerol (DMAG). The presence of DMAG was subsequently confirmed in other slow growing pathogenic species, including Mycobacterium tuberculosis. DMAG production was stimulated in the presence of exogenous glycerol. Interestingly, DMAG appears structurally identical to the terminal portion of the mycolylated arabinosyl motif of mAGP, and the metabolic relationship between these two components was provided using antitubercular drugs such as ethambutol or isoniazid known to inhibit the biosynthesis of arabinogalactan or mycolic acid, respectively. Finally, DMAG was identified in the cell wall of M. tuberculosis. This opens the possibility of a potent biological function for DMAG that may be important to mycobacterial pathogenesis.     

Evaluation of COMU as a coupling reagent for in situ neutralization Boc solid phase peptide synthesis

Benzotriazole‐based coupling reagents have dominated the last two decades of solid phase peptide synthesis. However, a growing interest in synthesizing complex peptides has stimulated the search for more efficient and low‐cost coupling reagents, such as COMU which has been introduced as a nonexplosive alternative to the classic benzotriazole coupling reagents. Here, we present a comparative study of the coupling efficiency of COMU with the benzotriazole‐based HBTU and HCTU for use in in situ neutralization Boc‐SPPS. Difficult sequences, such as ACP(65–74), Jung–Redeman 10‐mer, and HIV‐1 PR(81–99), were used as model target peptides on polystyrene‐based resins, as well as polyethylene glycol‐based resins. Coupling yields obtained using fast in situ Boc‐SPPS cycles were determined with the quantitative ninhydrin test as well as via LC‐MS analysis of the crude cleavage products. Our results demonstrate that COMU coupling efficiency was less effective compared to HBTU and HCTU with HCTU ≥ HBTU > COMU, when polystyrene‐based resins were employed. However, when the PEG resin was employed in combination with a safety catch amide (SCAL) linker, more comparable yields were observed for the three coupling reagents with the same ranking HCTU ≥ HBTU > COMU. Copyright © 2012 European Peptide Society and John Wiley & Sons, Ltd.