Cholesterol, lanosterol, and ergosterol attenuate the membrane association of LL-37(W27F) and temporin L

Sterols impart significant changes to the biophysical properties of lipid bilayers. In this regard the impact of cholesterol on membrane organization and dynamics is particularly well documented and serves for comparison with other sterols. However, the factors underlying the molecular evolution of cholesterol remain enigmatic. To this end, cholesterol attenuates membrane perturbation by the so-called antimicrobial peptides (AMPs), produced ubiquitously by eukaryotic cells to combat bacterial infections by compromising the permeability barrier function of the microbial target membranes. In the present study, we addressed the effects of cholesterol, ergosterol, and lanosterol on the membrane association of two structurally and functionally diverse AMPs viz. LL-37(F27W) and temporin L (TemL) using fluorescence spectroscopy. Interestingly, sterol concentration dependent effects on the membrane association of these peptides were observed. At X_Sterol = 0.5 cholesterol was most effective in reducing the membrane intercalation of both LL-37(F27W) and TemL, the corresponding efficiencies of the three sterols decreasing as cholesterol > lanosterol ≥ ergosterol, and cholesterol > lanosterol > ergosterol. It is conceivable that part of the selection pressure for the chemical evolution of cholesterol may have derived from the ability to protect the AMP-secreting host cell from the membrane damaging action of the antimicrobial peptides.     

CID-miRNA: a web server for prediction of novel miRNA precursors in human genome

microRNAs (miRNA) are a class of non-protein coding functional RNAs that are thought to regulate expression of target genes by direct interaction with mRNAs. miRNAs have been identified through both experimental and computational methods in a variety of eukaryotic organisms. Though these approaches have been partially successful, there is a need to develop more tools for detection of these RNAs as they are also thought to be present in abundance in many genomes. In this report we describe a tool and a web server, named CID-miRNA, for identification of miRNA precursors in a given DNA sequence, utilising secondary structure-based filtering systems and an algorithm based on stochastic context free grammar trained on human miRNAs. CID-miRNA analyses a given sequence using a web interface, for presence of putative miRNA precursors and the generated output lists all the potential regions that can form miRNA-like structures. It can also scan large genomic sequences for the presence of potential miRNA precursors in its stand-alone form. The web server can be accessed at http://mirna.jnu.ac.in/cidmirna/.     

Novel bacterial artificial chromosome vector pUvBBAC for use in studies of the functional genomics of Listeria spp

Bacterial artificial chromosome (BAC) vectors are important tools for microbial genome research. We constructed a novel BAC vector, pUvBBAC, for replication in both gram-negative and gram-positive bacterial hosts. The pUvBBAC vector was used to generate a BAC library for the facultative intracellular pathogen Listeria monocytogenes EGD-e. The library had insert sizes ranging from 68 to 178 kb. We identified two recombinant BACs from the L. monocytogenes pUvBBAC library that each contained the entire virulence gene cluster (vgc) of L. monocytogenes and transferred them to a nonpathogenic Listeria innocua strain. Recombinant L. innocua strains harboring pUvBBAC+vgc1 and pUvBBAC+vgc2 produced the vgc-specific listeriolysin (LLO) and actin assembly protein ActA and represent the first reported cloning of the vgc locus in its entirety. The use of the novel broad-host-range BAC vector pUvBBAC extends the versatility of this technology and provides a powerful platform for detailed functional genomics of gram-positive bacteria as well as its use in explorative functional metagenomics.     

Open‐Circuit Voltage in Organic Solar Cells: The Impacts of Donor Semicrystallinity and Coexistence of Multiple Interfacial Charge‐Transfer Bands

In organic solar cells (OSCs), the energy of the charge‐transfer (CT) complexes at the donor–acceptor interface, E _CT, determines the maximum open‐circuit voltage (V _OC). The coexistence of phases with different degrees of order in the donor or the acceptor, as in blends of semi‐crystalline donors and fullerenes in bulk heterojunction layers, influences the distribution of CT states and the V _OC enormously. Yet, the question of how structural heterogeneities alter CT states and the V _OC is seldom addressed systematically. In this work, we combine experimental measurements of vacuum‐deposited rubrene/C₆₀ bilayer OSCs, with varying microstructure and texture, with density functional theory calculations to determine how relative molecular orientations and extents of structural order influence E _CT and V _OC. We find that varying the microstructure of rubrene gives rise to CT bands with varying energies. The CT band that originates from crystalline rubrene lies up to ≈0.4 eV lower in energy compared to the one that arises from amorphous rubrene. These low‐lying CT states contribute strongly to V _OC losses and result mainly from hole delocalization in aggregated rubrene. This work points to the importance of realizing interfacial structural control that prevents the formation of low E _CT configurations and maximizes V _OC.     

Encapsulation of glucose oxidase and an oxygen-quenched fluorophore in polyelectrolyte-coated calcium alginate microspheres as optical glucose sensor systems

Microspheres coated with polyelectrolyte multilayers (PEM's) are being investigated for potential use as implantable biosensors-so-called "smart tattoos." In this work, the feasibility of this approach for glucose sensors was demonstrated by glucose oxidase encapsulated within calcium alginate microspheres, followed by entrapment of an oxygen-quenched ruthenium compound in the same microstructure. A novel feature of these microdevices is the formation of multilayer nanofilms on the surface of the microspheres, used to stabilize enzyme entrapment and control substrate diffusion. Confocal microscopy was used to confirm the stable encapsulation of sensor chemistry. The reversible response of sensors to step changes in glucose was observed, and preliminary experimental data were compared to theoretical predictions produced by a computational model. These findings demonstrate the promise of the described nanoengineering approach for production of functional implantable glucose sensor materials.     

Effects of remote mutation on the autolysis of HIV-1 PR: X-ray and NMR investigations

Autolysis rates of the C95M and C95M/C1095A mutants of a HIV-1 protease tethered dimer have been determined by real time NMR and it is observed that the double mutant has approximately two times higher rate. X-ray structure of the C95M/C1095A double mutant has been solved and refined to 2.1 A resolution. Comparison of the double mutant structure with that of C95M single mutant reveals that there is a shift in the position of the catalytic aspartates and the bound catalytic water. The mutation also causes a loss of hydrophobic packing near the dimerization domain of the protein. These observations demonstrate that subtle changes are adequate to cause significant changes in the rate of autolysis of the double mutant. This provides a rationale for the effects of remote mutations on the activity and drug resistance of the enzyme.     

Mining genomic databases to identify novel hydrogen producers

The realization that fossil fuel reserves are limited and their adverse effect on the environment has forced us to look into alternative sources of energy. Hydrogen is a strong contender as a future fuel. Biological hydrogen production ranges from 0.37 to 3.3 moles H(2) per mole of glucose and, considering the high theoretical values of production (4.0 moles H(2) per mole of glucose), it is worth exploring approaches to increase hydrogen yields. Screening the untapped microbial population is a promising possibility. Sequence analysis and pathway alignment of hydrogen metabolism in complete and incomplete genomes has led to the identification of potential hydrogen producers.