Bioluminescence characteristics of the fruiting body of Mycena chlorophos

Bioluminescent fungi are widely distributed on land and most belong to the class Basidomycetes. Light of about 530 nm wavelength maximum is emitted continuously. The molecular basis for the light‐emitting process remains unclear. We investigated the characteristics of the bioluminescence using cultivated fruiting bodies of M. chlorophos. Only fresh fruiting bodies exhibited long‐lasting light emission; rapid decay of light emission was observed with frozen and freeze‐dried samples. Freeze‐dried samples can be stored at room temperature under dry conditions and may be useful for the isolation of luciferin. The light emission of the fresh fruiting bodies was maintained in various buffers at varying pH; it could be stopped with pH 4 acetate buffer and could be recovered at pH 6. The isolation of luciferin from the fresh fruiting bodies might be possible by the control of buffer pH. The effect of temperature on the light emission of fruiting bodies indicated that bioluminescence in M. chlorophos may involve enzymatic reaction(s). The solubilization of bioluminescent components from the fruiting bodies could not be achieved with various surfactants. Copyright © 2011 John Wiley & Sons, Ltd.     

Characterization of Switch Phenotypes in <Emphasis Type="Italic">Candida albicans</Emphasis> Biofilms

The aim of this study was to characterize switch phenotypes in Candida albicans biofilms. Cells of Candida albicans 192887g biofilms (24 h) were resuspended and these together with their planktonic counterparts were separately inoculated on Lee’s medium agar supplemented with arginine and zinc, at 25 °C for 9 days, for colony formation. The different switch phenotypes, as reflected by varying colony morphologies, were then examined for their (i) stability under various growth conditions, (ii) carbohydrate assimilation profiles, (iii) susceptibility to the polyene antifungal, nystatin, (iv) adhering and biofilm-forming ability, (v) filamentation, and (vi) growth rate in yeast nitrogen base medium supplemented with 100 mM glucose. Our data showed that the frequency of phenotypic switching in C. albicans biofilms was approximately 1%. Compared with the planktonic yeasts, cells derived from candidal biofilms generated one of the phenotypes less frequently (Chi-square-tests: P = 0.017). The five phenotypes derived from the biofilm growth demonstrated differing profiles for carbohydrate assimilation, adhesion, biofilm formation, filamentation, and growth rate. These findings reported here, for the first time, imply that phenotypic switching in the candidal biofilms differs from that in the planktonic growth, and affects multiple biological attributes.     

Tuning the biological properties of amphipathic alpha-helical antimicrobial peptides: rational use of minimal amino acid substitutions

In nature, alpha-helical antimicrobial peptides present the small and flexible residue glycine at positions 7 or 14 with a significant frequency. Based on the sequence of the non-proteinogenic alpha-helical model peptide P1(Aib7), with a potent, broad spectrum antimicrobial activity, six peptides were designed by effecting a single amino acid substitution to investigate how tuning the structural characteristics at position 7 could lead to optimization of selectivity without affecting antimicrobial activity against a broad panel of multidrug resistant bacterial and yeast indicator strains. The relationship between structural features (size/hydrophobicity of the side chain as well as conformation and flexibility) and biological activity, in terms of minimum inhibitory concentration, membrane permeabilization kinetics and lysis of red blood cells are discussed. On conversion of the peptide to proteinogenic residues, these principles allowed development of a potent antimicrobial peptide with a reduced cytotoxicity. However, while results suggest that both hydrophobicity of residue 7 and chain flexibility at this position can be modulated to improve selectivity, position 14 is less tolerant of substitutions.     

Duplications between direct repeats stabilized by DNA secondary structure occur preferentially in the leading strand during DNA replication

To ascertain a leading or lagging strand preference for duplication mutations, several short DNA sequences, i.e. mutation inserts, were designed that should demonstrate an asymmetric propensity for duplication mutations in the two complementary DNA strands during replication. The design of the mutation insert involved a 7-bp quasi inverted repeat that forms a remarkably stable hairpin in one DNA strand, but not the other. The inverted repeat is asymmetrically placed between flanking direct repeats. This sequence was cloned into a modified chloramphenicol acetyltransferase (CAT) gene containing a −1 frameshift mutation. Duplication of the mutation insert restores the reading frame of the CAT gene resulting in a chloramphenicol resistant phenotype. The mutation insert showed greater than a 200-fold preference for duplication mutations during leading strand, compared with lagging strand, replication. This result suggests that misalignment stabilized by DNA secondary structure, leading to duplication between direct repeats, occurred preferentially during leading strand synthesis.     

Differential effect of IL-4 and IL-13 on the expression of recombination-activating genes in mature B cells from human peripheral blood

We examined the expression of recombination-activating genes (RAG-1 and RAG-2) and activation-induced cytidine deaminase (AID) by mature human blood B cells stimulated with anti-CD40 in the presence of IL-4 or IL-13. IL-4 was an effective cofactor for RAG-1 and RAG-2 expression, whereas IL-13 was not. In addition, IL-4-dependent RAG expression combined with AID and IgE expression allowed predominant expression of newly rearranged lambda light chains on IgE+ cells generated from kappa+ cells. Although the magnitudes of IL-4- and IL-13-dependent AID and IgE expression were related to expression levels of binding subunits of the IL-4 and IL-13 receptors, IL-13 was ineffective for light chain replacement in the induced IgE+ cells due to the failure in RAG expression. Our studies using mature blood B cells indicate that IL-4-responsive cells, unlike IL-13-responsive cells, undergo lambda gene rearrangement leading to replacement in parallel with RAG expression and suggest that this replacement may contribute to the regulation of affinity maturation of IgE antibodies.     

Mating type-dependent constraints on the mobility of the left arm of yeast chromosome III

Mating-type gene (MAT) switching in budding yeast exhibits donor preference. MATa preferentially recombines with HML near the left telomere of chromosome III, whereas MATalpha prefers HMR near the right telomere. Donor preference is controlled by the recombination enhancer (RE) located proximal to HML. To test if HML is constrained in pairing with MATalpha, we examined live-cell mobility of LacI-GFP-bound lactose operator (lacO) arrays inserted at different chromosomal sites. Without induction of recombination, lacO sequences adjacent to HML are strongly constrained in both MATalpha and RE-deleted MATa strains, compared with MATa. In contrast, chromosome movement at HMR or near a telomere of chromosome V is mating-type independent. HML is more constrained in MATa Deltare and less constrained in MATa RE+ compared with other sites. Although HML and MATa are not prealigned before inducing recombination, the three-dimensional configuration of MAT, HML, and HMR is mating-type dependent. These data suggest there is constitutive tethering of HML, which is relieved in MATa cells through the action of RE.     

Protein Folding Requires Crowd Control in a Simulated Cell

Macromolecular crowding has a profound effect upon biochemical processes in the cell. We have computationally studied the effect of crowding upon protein folding for 12 small domains in a simulated cell using a coarse-grained protein model, which is based upon Langevin dynamics, designed to unify the often disjoint goals of protein folding simulation and structure prediction. The model can make predictions of native conformation with accuracy comparable with that of the best current template-free models. It is fast enough to enable a more extensive analysis of crowding than previously attempted, studying several proteins at many crowding levels and further random repetitions designed to more closely approximate the ensemble of conformations. We found that when crowding approaches 40% excluded volume, the maximum level found in the cell, proteins fold to fewer native-like states. Notably, when crowding is increased beyond this level, there is a sudden failure of protein folding: proteins fix upon a structure more quickly and become trapped in extended conformations. These results suggest that the ability of small protein domains to fold without the help of chaperones may be an important factor in limiting the degree of macromolecular crowding in the cell. Here, we discuss the possible implications regarding the relationship between protein expression level, protein size, chaperone activity and aggregation.     

Synthesis, structure and photocatalytic activity of a remarkably bent, cofacial ethene-linked diiron (III) μ-oxobisporphyrin

Synthesis and characterization of cis and trans 1,2-bis[Chloroiron(III) 5-(2,3,7,8,12,13,17,18-octaethylporphyrinyl)]ethenes are described. Upon treatment with 5% NaOH, cis form of the molecule immediately converts to remarkably bent diiron (III)-μ-oxo bisporphyrin which transforms to cis bisChloroFe(III)porphyrin again by the addition of 5% HCl. This facile transformation is reversible with sharp change in color in which the bisporphyrin platform ‘open’ and ‘close’ its binding pockets with very high vertical flexibility in a single molecular framework. Single crystal X-ray structural characterization reveals cis diiron(III)-μ-oxo bisporphyrin in which Fe-O-Fe unit is remarkable bent with 150.9(2)° angle. Two porphyrin rings in the molecule are not slipped but face-to-face in a fully eclipsed geometry and are placed so close that some of the carbon atoms from each of the macrocycles are driven to be essentially less than the van der Waals contacts (<3.4 Å). Two rings in the oxo-bridged dimer also make the interplanar angle of 27.7° instead of expected angle of 60° due to the bridging alkenic bond. EPR, ¹H NMR and Mössbauer spectral data are indicative of strong anti-ferromagnetic coupling between two high-spin iron(III) centers via bridging oxo group. The complex catalyzes the rapid photoinduced oxygenation of phosphites under mild condition using aerial O₂. Electrochemical data reveals that the diiron(III)-μ-oxo bisporphyrin in dichloromethane undergoes four reversible/quasi-reversible one electron oxidation and one electron reductions. The presence of two porphyrin macrocycles within a short distance in the μ-oxo species makes the porphyrin core highly nonplanar and more electron rich that might responsible for easier oxidations compared to [Fe(OEP)]₂O.     

The Spatial Structure of Variability in a Semi-arid, Fluvial Ecosystem

The arrangement and composition of flowpath types within a given network are thought to govern its functioning. This concept assumes that different flowpath types are functionally distinct. We investigated this assumption in a fluvial ecosystem by comparing the riparian zone, parafluvial zone (in-channel gravel bars), and surface stream. We hypothesized that differences in advection, uptake, and sorption would render material cycles more (a) open and (b) mutable in the surface stream, whereas the converse would occur in the riparian zone, and an intermediate state would be seen in the intervening parafluvial zone. To test our first hypothesis, we predicted that spatial heterogeneity in solute concentrations would be least in the surface stream, greater in the parafluvial zone, and greatest in the riparian zone. Using a null model, we ascertained that this pattern was shown by all solute species we examined (nitrate, ammonium, total dissolved inorganic nitrogen [DIN], dissolved organic N, total dissolved N, soluble reactive phosphorus, dissolved organic carbon, and chloride). To test our second hypothesis, we predicted that temporal change in spatial heterogeneity would be greatest in the surface stream, less in the parafluvial zone, and least in the riparian zone. Nitrate, DIN, and chloride showed this pattern. In particular, surface stream inorganic N was less spatially variable following months of high rainfall. According to an extant hypothesis, these results suggest that inorganic N processing may be a stable function in this ecosystem. Other solute species did not support our second prediction, perhaps because their retention and release dynamics are influenced principally by geochemistry. Generally, our findings indicate that a geomorphic template can generate spatial patterns in ecosystem function, warranting an expansion of the spiraling framework to a variety of flowpath types.     

Asparagales Telomerases which Synthesize the Human Type of Telomeres

The order of monocotyledonous plants Asparagales is attractive for studies of telomere evolution as it includes three phylogenetically distinct groups with telomeres composed of TTTAGGG (Arabidopsis-type), TTAGGG (human-type) and unknown alternative sequences, respectively. To analyze the molecular causes of these switches in telomere sequence (synthesis), genes coding for the catalytic telomerase subunit (TERT) of representative species in the first two groups have been cloned. Multiple alignments of the sequences, together with other TERT sequences in databases, suggested candidate amino acid substitutions grouped in the Asparagales TERT synthesizing the human-type repeat that could have contributed to the changed telomere sequence. Among these, mutations in the C motif are of special interest due to its functional importance in TERT. Furthermore, two different modes of initial elongation of the substrate primer were observed in Asparagales telomerases producing human-like repeats, which could be attributed to interactions between the telomerase RNA subunit (TR) and the substrate. Electronic Supplementary Material Supplementary material is available for this article at and is accessible for authorized users.