Structural and chemical characterization of S-layers of selected strains ofBacillus stearothermophilus andDesulfotomaculum nigrificans

The structures, amino acid- and neutral sugar compositions of the crystalline surface layers (S-layers) of four selected strains each ofBacillus stearothermophilus andDesulfotomaculum nigrificans were compared. Among the four strains of each species a remarkable diversity in the molecular weights of the S-layer subunits and in the geometry and constants of the S-layer lattices was apparent. The crystalline arrays included hexagonal (p6), square (p4) and oblique (p2) lattices. In vitro self-assembly of isolated S-layer subunits (or S-layer fragments) led to the formation of flat sheets or open-ended cylindrical assembly products. The amino acid composition of the S-layers exhibited great similarities and was predominantly acidic. With the exception of the S-layers of two strains ofB. stearothermophilus (where only traces of neutral sugars could be detected), all other S-layer proteins seemed to be glycosylated. Among these strains significant differences in the amount and composition of the glycan portions were found. Based on this diversity interesting questions may be asked about the biological significance of the carbohydrate units of glycoproteins in prokaryotic organisms.     

Interactions of Meso-tetra-(4-N-oxyethylpyridyl) Porphyrin,Its 3-N Analog and Their Metallocomplexes with Duplex DNA

Abstract

Interactions of meso-tetra-(4-N-oxyethylpyridyl) porphyrin (TOEPyP(4)), its 3-N analog (TOEPyP(3)) and their Co, Cu, Ni, Zn metallocomplexes with duplex DNA have been investigated by uv/visible absorbance and circular dichrosim spectroscopies. Results reveal the interactions of these complexes with duplex DNA are of two types. (1) External binding of duplex DNA by metalloporphyrins containing Zn and Co, and (2) Binding of duplex DNA both externally and internally (by intercalation) by porphyrins not containing metals, and metalloporphyrins containing Cu and Ni. Results indicate that (4N-oxyethylpyridyl) porphyrins intercalate more preferably in the structure of duplex DNA and have weaker external binding than 3N-porphyrins.     

Mice with deletion of the mitochondrial glycerol-3-phosphate dehydrogenase gene exhibit a thrifty phenotype: effect of gender

To define the role of mitochondrial glycerol-3-phosphate dehydrogenase (mGPD; EC 1.1.99.5) in energy balance and intermediary metabolism, we studied transgenic mice not expressing mGPD (mGPD-/-). These mice had approximately 14% lower blood glucose; approximately 50% higher serum glycerol; approximately 80% higher serum triglycerides; and at thermoneutrality, their energy expenditure (Qo(2)) was 15% lower than in wild-type (WT) mice. Glycerol-3-phosphate levels and lactate-to-pyruvate ratios were threefold elevated in muscle, but not in liver, of mGPD-/- mice. WT and mGPD-/- mice were then challenged with a high-fat diet, fasting, or food restriction. The high-fat diet caused more weight gain and adiposity in mGPD-/- than in WT female mice, without the genotype differentially affecting Qo(2) or energy intake. After a 30-h fast, WT female lost 60% more weight than mGPD-/- mice but these latter became more hypothermic. When energy intake was restricted to 50-70% of the ad libitum intake for 10 days, mGPD-/- female mice lost less weight than WT controls, but they had lower Qo(2) and body temperature. WT and mGPD-/- male mice did not differ significantly in their responses to these challenges. These results show that the lack of mGPD causes significant alterations of intermediary metabolism, which are more pronounced in muscle than liver and lead to a thrifty phenotype that is more marked in females than males. Lower T(4)-to-T(3) conversion in mGPD-/- females and a greater reliance of normal females on mGPD to respond to high-fat diets make the lack of the enzyme more consequential in the female gender.     

Multivariate approach for selecting sets of differentially expressed genes

An important problem addressed using cDNA microarray data is the detection of genes differentially expressed in two tissues of interest. Currently used approaches ignore the multidimensional structure of the data. However it is well known that correlation among covariates can enhance the ability to detect less pronounced differences. We use the Mahalanobis distance between vectors of gene expressions as a criterion for simultaneously comparing a set of genes and develop an algorithm for maximizing it. To overcome the problem of instability of covariance matrices we propose a new method of combining data from small-scale random search experiments. We show that by utilizing the correlation structure the multivariate method, in addition to the genes found by the one-dimensional criteria, finds genes whose differential expression is not detectable marginally.     

Understanding the structural requirements of 4-anilidopiperidine analogues for biological activities at mu and delta opioid receptors

New 4-anilidopiperidine analogues in which the phenethyl group of fentanyl was replaced by several aromatic ring-contained amino acids (or acids) were synthesized to study the biological effect of the substituents on mu and delta opioid receptor interactions. These analogues showed broad (47 nM-76 microM) but selective (up to 17-fold) binding affinities at the mu opioid receptor over the delta opioid receptor, as predicted from the message-address concept.     

Interactive effects of chromate and arsenate on their uptake and speciation in Pteris ensiformis

Background and aims

Arsenate (AsV) and chromate (CrVI) inhibit each other’s uptake and translocation in As-hyperaccumulator Pteris vittata. In the present study, we extended the research to As-sensitive plant Pteris ensiformis to better understand the mechanism of their interactions.

Methods

Plants were exposed to 0, 0.75 or 7.5 mg L^?1 AsV and 0, 0.52, or 5.2 mg L^?1 CrVI for 7 d in hydroponics. Arsenic and Cr speciation were determined in nutrient solutions and plant biomass.

Results

P. ensiformis accumulated high levels of As and Cr in the rhizomes and roots with low levels in the fronds. However, P. ensiformis was much more effective in taking up Cr than As, as much more Cr was accumulated in the roots (306–6015 vs. 87–642 mg kg^?1). AsV and CrVI increased each other’s uptake in the rhizomes and roots when co-present. The AsV and CrVI taken up by P. ensiformis were reduced to arsenite (AsIII) and chromite (CrIII), possibly serving as detoxification mechanism.

Conclusions

Uptake of As and Cr induced oxidative stress as indicated by increased lipid peroxidation and electrical conductivity. Arsenic and Cr increased each other’s uptake by P. ensiformis.

     

TMEFF2 shedding is regulated by oxidative stress and mediated by ADAMs and transmembrane serine proteases implicated in prostate cancer

TMEFF2 is a type I transmembrane protein with two follistatin (FS) and one EGF‐like domain over‐expressed in prostate cancer; however its biological role in prostate cancer development and progression remains unclear, which may, at least in part, be explained by its proteolytic processing. The extracellular part of TMEFF2 (TMEFF2‐ECD) is cleaved by ADAM17 and the membrane‐retained fragment is further processed by the gamma‐secretase complex. TMEFF2 shedding is increased with cell crowding, a condition associated with the tumour microenvironment, which was mediated by oxidative stress signalling, requiring jun‐kinase (JNK) activation. Moreover, we have identified that TMEFF2 is also a novel substrate for other proteases implicated in prostate cancer, including two ADAMs (ADAM9 and ADAM12) and the type II transmembrane serine proteinases (TTSPs) matriptase‐1 and hepsin. Whereas cleavage by ADAM9 and ADAM12 generates previously identified TMEFF2‐ECD, proteolytic processing by matriptase‐1 and hepsin produced TMEFF2 fragments, composed of TMEFF2‐ECD or FS and/or EGF‐like domains as well as novel membrane retained fragments. Differential TMEFF2 processing from a single transmembrane protein may be a general mechanism to modulate transmembrane protein levels and domains, dependent on the repertoire of ADAMs or TTSPs expressed by the target cell.     

Temporal and transient expression of olive enoyl-ACP reductase gene during flower and fruit development.

Enoyl-ACP reductase is a catalytic component of the fatty acid synthetase (FAS) type II system in plants that is involved in the de novo fatty acid biosynthesis in plastids. A cDNA encoding an enoyl-ACP reductase responsible for the removal of the trans-unsaturated double bonds to form saturated acyl-ACP has been isolated from a library made from ripening fruits of Olea europaea L. The predicted protein contains 393 amino acid residues including a consensus chloroplast specific transit peptide. A strong homology was observed when olive enoyl-ACP reductase aligned with other plant sequences. Southern hybridization analysis revealed that enoyl-ACP reductase is encoded by a single gene in olives. Northern hybridization showed a transient expression of the enoyl-ACP reductase (ENR) gene at early stages of drupe (5-7 weeks after flowering, WAF), embryo and endosperm (13-16 WAF) while in mesocarp (13-19 WAF) the expression remained at high levels. In situ hybridization showed particularly prominent expression in the palisade and vascular tissue of young leaves, the tapetum, developing pollen grains and vascular tissue of anthers and to less extent in the embryo sac and transmitting tissue of the carpel. The distinctive spatial and temporal regulation of the ENR gene is consistent with major roles, not only in thylakoid membrane formation and fatty acid deposition, but also in the provision of precursor molecules for the biosynthesis of oxilipins that are important in plant tissues involved in transportation and reproduction.     

Isolation,purification and physicochemical characterization of water‐soluble Bacillus thuringiensis melanin

Melanins are widely used in medicine, pharmacology, cosmetics and other fields. Although several technologies for the purification of water‐insoluble dioxyphenylalanine (DOPA) melanins have been described, a source of water‐soluble melanin is highly desirable. Here we describe an effective procedure for the isolation and purification of water‐soluble melanin using the culture medium of Bacillus thuringiensis subsp. galleriae strain K1. Water‐soluble melanin from this organism has an isoelectric point (pI = 3.0–3.2) and was purified optimally by adsorbtion using the IA‐1r resin and elution as a concentrated solution. The purified melanin obtained exhibited a similar infra‐red absorbtion spectrum to synthetic melanin and contained quinolic and phenolic structures and an amino acid content of around 20% after acid hydrolysis. The molecular weight of the purified melanin determined by SDS‐PAGE was 4 kDa and the electromagnetic spin resonance spectrum of the purified microbial melanin was a slightly asymmetric singlet without hyperfine structure with about 7 Gauss width of the line between points of the maximum incline and g = 2.006. The concentration of paramagnetic centers in melanin is 0.21 × 10¹⁸ spin/g. The results obtained provide a rapid, simple and inexpensive method for the large scale purification of water soluble melanin that may have widespread applications.