How might you compare mitochondria from different tissues and different species?

Mitochondria were isolated from the liver, kidney and mixed hindlimb skeletal muscle of three vertebrate species; the laboratory rat Rattus norvegicus, the bearded dragon lizard Pogona vitticeps, and the cane toad Bufo marinus. These vertebrate species are approximately the same body mass and have similar body temperatures. The content of cytochromes B, C, C1, and A were measured in these isolated mitochondria by oxidised–reduced difference spectra. Adenine nucleotide translocase (ANT) was measured by titration of mitochondrial respiration with carboxyactractyloside and the protein and phospholipid content of isolated mitochondria were also measured. Fatty acid composition of mitochondrial phospholipids was measured. Mitochondrial respiration was measured at 37°C under states III and IV conditions as well as during oligomycin inhibition. Species differed in the ratios of different mitochondrial cytochromes. Muscle mitochondria differed from kidney and liver mitochondria by having a higher ANT content relative to cytochrome content. Respiration rates were compared relative to a number of denominators and found to be most variable when expressed relative to mitochondrial protein content and least variable when expressed relative to mitochondrial cytochrome A and ANT content. The turnover of cytochromes was calculated and found to vary between 1 and 94 electrons s⁻¹. The molecular activity of mitochondrial cytochromes was found to be significantly positively correlated with the relative polyunsaturation of mitochondrial membrane lipids.     

Why is SCA12 different from other SCAs?

Spinocerebellar ataxia type 12 (SCA12), now described in European-American and Asian (Indian) pedigrees, is unique among the SCAs from clinical, pathological, and molecular perspectives. Clinically, the distinguishing feature is early and prominent action tremor with variability in other signs. Pathologically, brain MRIs also suggest variability, with prominent cortical as well as cerebellar atrophy. Genetically, SCA12 is caused by a CAG repeat expansion that does not encode polyglutamine; we speculate that the mutation may affect expression of the gene PPP2R2B, which encodes a brain-specific regulatory subunit of the protein phosphatase PP2A.     

Why are individuals so different from each other?

An important contributor to the differences between individuals derives from their plasticity. Such plasticity is widespread in organisms from the simple to the most complex. Adaptability plasticity enables the organism to cope with a novel challenge not previously encountered by its ancestors. Conditional plasticity appears to have evolved from repeated challenges from the environment so that the organism responds in a particular manner to the environment in which it finds itself. The resulting phenotypic variation can be triggered during development in a variety of ways, some mediated through the parent''s phenotype. Sometimes the organism copes in suboptimal conditions trading off reproductive success against survival. Whatever the adaptedness of the phenotype, each of the many types of plasticity demonstrates how a given genotype will express itself differently in different environmental conditions—a field of biology referred to as the study of epigenetics. The ways in which epigenetic mechanisms may have evolved are discussed, as are the potential impacts on the evolution of their descendants.     

Poly-β-hydroxybutyrate accumulation in bacterial consortia from different environments

The aim of the present study was to examine soil samples from various vegetation zones in terms of physicochemical properties, microbial communities, and isolation and identification (by polymerase chain reaction and transmission electron microscopy) of bacteria producing poly-β-hydroxybutyrates (PHBs). Soil samples were analysed originating from zones with heterogeneous environmental conditions from the Romanian Carpathian Mountains (mountain zone with alpine meadow, karstic zone with limestone meadow, hill zone with xerophilous meadow, and flood plain zone with hygrophilic meadow). Different bacterial groups involved in the nitrogen cycle (aerobic mesophilic heterotrophs, ammonifiers, denitrifiers, nitrifiers, and free nitrogen-fixing bacteria from Azotobacter genus) were analysed. Soil biological quality was assessed by the bacterial indicator of soil quality, which varied between 4.3 and 4.7. A colony polymerase chain reaction technique was used for screening PHB producers. With different primers, specific bands were obtained in all the soil samples. Some wild types of Azotobacter species were isolated from the 4 studied sites. Biodegradable polymers of PHB were assessed by negative staining in transmission electron microscopy. The maximum PHB granules density was obtained in the strains isolated from the xerophilous meadow (10-18 granules/cell), which was the most stressful environment from all the studied sites, as the physicochemical and microbiological tests proved.     

Production and characterization of β-glucosidases from different Aspergillus strains

-D-Glucosidase enzymes (-D-glucoside glucohydrolase, EC 3.2.1.21) from different Aspergillus strains (Aspergillus phoenicis, A. niger and A. carbonarius) were examined with respect to the enzyme production of the different strains using different carbon sources and to the effect of the pH and temperature on the enzyme activity and stability. An efficient and rapid purification procedure was used for purifying the enzymes. Kinetic experiments were carried out using p-nitrophenyl -D-glucopyranoside (pNPG) and cellobiose as substrates. Two different fermentation methods were employed in which the carbon source was glucose or wheat bran. Aspergillus carbonarius proved to be the less effective strain in -glucosidase production. Aspergillus phoenicis produced the highest amount of -glucosidase on glucose as carbon source however on wheat bran A. niger was the best enzyme producer. Each Aspergillus strain produced one single acidic -glucosidase with pI values in the range of pH 3.52–4.2. There was no significant difference considering the effect of the pH and temperature on the activity and stability among the enzymes from different origins. The enzymes examined have only -glucosidase activity. The kinetic parameters showed that all enzymes hydrolysed pNPG with higher efficiency than cellobiose. This shows that hydrophobic interaction plays an important role in substrate binding. The kinetic parameters demonstrated that there was no significant difference among the enzymes from different origins in hydrolysing pNPG and cellobiose as the substrates.     

Are data from different gene expression microarray platforms comparable?

Many commercial and custom-made microarray formats are routinely used for large-scale gene expression surveys. Here, we sought to determine the level of concordance between microarray platforms by analyzing breast cancer cell lines with in situ synthesized oligonucleotide arrays (Affymetrix HG-U95v2), commercial cDNA microarrays (Agilent Human 1 cDNA), and custom-made cDNA microarrays from a sequence-validated 13K cDNA library. Gene expression data from the commercial platforms showed good correlations across the experiments (r = 0.78-0.86), whereas the correlations between the custom-made and either of the two commercial platforms were lower (r = 0.62-0.76). Discrepant findings were due to clone errors on the custom-made microarrays, old annotations, or unknown causes. Even within platform, there can be several ways to analyze data that may influence the correlation between platforms. Our results indicate that combining data from different microarray platforms is not straightforward. Variability of the data represents a challenge for developing future diagnostic applications of microarrays.     

β-1,4-endoglucanases from Talaromyces amestolkiae: Production of glucooligosaccharides from different β-glucans

Abstract

This article presents the purification and characterization of two β-1,4-endoglucanases from Talaromyces amestolkiae. The cellulase activities secreted by this fungus were studied in the presence of different carbon sources, attaining the maximal levels in the presence of Avicel as carbon source. In these conditions, two glycosylated β-1,4-endoglucanases with molecular masses of 25,573?kDa (EG1) and 51,825?kDa (EG2), were purified. Both isoenzymes have acidic isoelectric points, 5.4 and 4.6, respectively. Their optimum pH and temperature, either in crudes or after purification, were in the range normally used for the simultaneous saccharification and fermentation in bioethanol production. In addition, the enzymatic hydrolysis of different β-glucans by both enzymes was studied. In the assayed conditions, both enzymes hydrolysed carboxymethylcellulose, a typical substrate for endoglucanases, although EG2 was much more efficient. However, EG1 was also able to hydrolyse lichenan and laminarin. These findings suggest the potential interest of EG2 for specific hydrolysis of cellulose, present in plant cell walls, to produce bioethanol, while the more promiscuous enzyme EG1 could be used for production of glucooligosaccharides.     

α-Fucosidases with different substrate specificities from two species of Fusarium

Two fungal-secreted α-fucosidases and their genes were characterized. FoFCO1 was purified from culture filtrates of Fusarium oxysporum strain 0685 grown on l-fucose and its encoding gene identified in the sequenced genome of strain 4287. FoFCO1 was active on p-nitrophenyl-α-fucoside (pNP-Fuc), but did not defucosylate a nonasaccharide (XXFG) fragment of pea xyloglucan. A putative α-fucosidase gene (FgFCO1) from Fusarium graminearum was expressed in Pichia pastoris. FgFCO1 was ～1,800 times less active on pNP-Fuc than FoFCO1, but was able to defucosylate the XXFG nonasaccharide. Although FgFCO1 and FoFCO1 both belong to Glycosyl Hydrolase family 29, they share <25 % overall amino acid identity. Alignment of all available fungal orthologs of FoFCO1 and FgFCO1 indicated that these two proteins belong to two subfamilies of fungal GH29 α-fucosidases. Fungal orthologs of subfamily 1 (to which FoFCO1 belongs) are taxonomically more widely distributed than subfamily 2 (FgFCO1), but neither was universally present in the sequenced fungal genomes. Trichoderma reesei and most species of Aspergillus lack genes for either GH29 subfamily.     

Are Antarctic suspension-feeding communities different from those elsewhere in the world?

This paper reviews the trophic ecology of benthic suspension feeders in Antarctic shelf communities, studied within SCAR's EASIZ Programme, in comparison with published information from other seas. Dense benthic suspension-feeder communities capture large quantities of particles and may directly regulate primary, and indirectly, secondary production in littoral food chains. Most work has been performed in temperate and tropical seas; however, little is known about suspension feeders in cold environments. Recent studies on Antarctic littoral benthic suspension feeders suggest the period of winter inactivity may last only a few weeks. This contrasts with the hypothesis that in Antarctic communities there is a prolonged period of minimal activity lasting at least 6 months during the austral winter. Results from other oceans may explain how dense benthic communities could develop under such conditions. Alternative food sources, i.e. the "fine fraction", sediment resuspension, lateral advection and efficient food assimilation may play a significant role in the development of suspension-feeder dominated, very diversified, high biomass and three-dimensionally structured communities on the Antarctic shelf.     

Analysis of human extrachromosomal DNA elements originating from different β-satellite subfamilies

By screening total human DNA with probes derived from the small polydisperse circular (spc) DNA fraction of cultured human cells, we identified three clones that carry long stretches of -satellite DNA. Further experiments have shown that the three sequences belong to at least two different -satellite subfamilies, which are characterized by different higher order subunits. Members of one of these subfamilies are located in the cytological satellites of all acrocentric chromosomes, whereas members of another are located on the short arms of the acrocentrics on both sides of the stalk regions and also in the centromeric regions of chromosomes 1 and 9. This is the first time that -satellite sequences obtained from the spcDNA of human cells have been assigned to -satellite subfamilies that are organized as long arrays of tandemly arranged higher order monomers. This indicates that -satellite sequences can be excised from their chromosomal loci via intrastrand-recombination processes.This paper is dedicated to Prof. Dr. Ulrich Wolf on his 60th birthday, January 1993     

Is population subdivision different from speciation? From phylogeography to species delimitation

Species‐level diversity and the underlying mechanisms that lead to the formation of new species, that is, speciation, have often been confounded with intraspecific diversity and population subdivision. The delineation between intraspecific and interspecific divergence processes has received much less attention than species delimitation. The ramifications of confounding speciation and population subdivision are that the term speciation has been used to describe many different biological divergence processes, rendering the results, or inferences, between studies incomparable. Phylogeographic studies have advanced our understanding of how spatial variation in the pattern of biodiversity can begin, become structured, and persist through time. Studies of species delimitation have further provided statistical and model‐based approaches to determine the phylogeographic entities that merit species status. However, without a proper understanding and delineation between the processes that generate and maintain intraspecific and interspecific diversity in a study system, the delimitation of species may still not be biologically and evolutionarily relevant. I argue that variation in the continuity of the divergence process among biological systems could be a key factor leading to the enduring contention in delineating divergence patterns, or species delimitation, meriting future comparative studies to help us better understand the nature of biological species.     

Is Fusarium culmorum isotrichodermin-15-hydroxylase different from other fungal species?

Fusarium spp. are ubiquitous fungi infecting cereals and grains, and therefore constitute a major problem for agriculture. Their trichothecene metabolites, and in particular deoxynivalenol and its 3-acetylated derivative, are the mycotoxins involved. The major metabolite produced by Fusarium culmorum is 3-acetyldeoxynivalenol. Studies in vivo with Fusarium culmorum have established that its tricyclic intermediate, isotrichodermin, is a major biosynthetic precursor, which is hydroxylated at position 15 to give 15-deacetylcalonectrin, prior to being converted to the product. In a preliminary in vitro investigation of the cell-free system involved in this transformation, we suggested that cytochrome P450 enzymes are not involved. In this paper, the isotrichodermin-15-hydroxylase from the microsomal fraction of Fusarium culmorum was solubilized and partially purified (60 fold). Our studies with cofactors indicate that this enzyme is a flavoprotein, and the inducers tested highly indicate that indeed the hydroxylase is not attached to cytochrome P450. This is particularly interesting, since the only other enzyme catalyzing the same reaction isolated from Fusarium sporotrichiodes is attached to cytochrome P450.     

Purification and properties of two different α1-protease inhibitors from mouse plasma

Two similar but distinct forms of α1-protease inhibitor (α1-PI) have been isolated and purified 120-fold to homogeneity from the plasma of female, white Swiss (Ha/ICR) mice. The two inhibitors can be separated by chromatography on DEAE-cellulose using a shallow NaCl gradient at pH 8.9 for elution. Because of their differing specificities for elastase and trypsin we have labeled the two inhibitors α1-PI(E) and α1-PI(T), respectively. The apparent M_r for both proteins, as estimated by gel exclusion chromatography, is approximately 53,000 daltons. However by polyacrylamide gel electrophoresis in the presence of SDS, α1-PI(T) has an apparent m_r of 65,000 while the apparent m_r of α1-PI(E) is 55,000. These results suggest differences in charge and carbohydrate composition. The two mouse inhibitors also have different AT-terminal amino acids. Like human α1-PI the mouse inhibitors form stable complexes with proteases. However they differed from human α1-PI in that they were not found to neutralize either human thrombin or plasmin. While α1-PI(E) inhibits bovine pancreatic trypsin, chymotrypsin, and porcine pancreatic elastase, α1-PI(T) is an effective inhibitor only of trypsin. Plasma levels of α1-PI(E) increase significantly 24 h after stimulation of the acute phase reaction while those of α1-PI(T) do not. Our data suggest that α1-PI(E) and α1-PI(T) are products of different genes.     

Comparison of rhizobitoxine-induced inhibition of β-cystathionase from different bradyrhizobia and soybean genotypes

The enzyme -cystathionase catalyzes the conversion of cystathionine to homocysteine in both plants and bacteria. Preparations of this enzyme taken from both Salmonella and spinach (Spinacia oleracea L.) have been shown to be irreversibly inhibited by low concentrations of rhizobitoxine (RT), a chlorosis-inducing phytotoxin produced by some strains of soybean bradyrhizobia. The sensitivities of -cystathionase from bradyrhizobia and soybean are not well characterized. Therefore, we purified -cystathionase from selected bradyrhizobia and soybean genotypes that have been shown to exhibit differences in RT production and apparent RT sensitivities, respectively. Enzyme purified from E. coli strain DH52 was used for comparison. The enzymes differed in their physiological properties and RT sensitivities. Overall, the -cystathionase enzymes purified from bradyrhizobia were more sensitive to RT than were those from the soybean cultivars. Kinetic studies showed that the nature of the RT-induced inhibition also differed between the two sources. The enzymes from bradyrhizobia exhibited inhibition that was [RT]-dependent, whereas the enzymes from soybean showed a time-dependent inhibition. These contrasting characteristics may in part reflect differences in active site accessibility, amino acid components, and associated RT diffusion rates. However, in all cases the inhibition caused by RT showed a typical substrate-competitive inhibition pattern.Abbreviations RT rhizobitoxine - PLP pyridoxal phosphate - DTT dithioerythritol - PMSF phenylmethylsulfonyl fluoride - HTP hydroxyapatite Published as Paper No. 1571 in the Journal Series of the Delaware Agricultural Experiment Station.     

The Bemisia tabaci species complex： Additions from different parts of the world

Bemisia tabaci is one of the most threatening pests in many crops. We sequenced part of the mitochondrial cytochrome oxidase I gene from fifty whitefly populations collected in Indonesia, Thailand, India and China. Nineteen unique sequences （haplotypes） of the cytochrome oxidase I were identified in these populations. They were combined with sequences available in databases, resulting in a total of 407 haplotypes and analyzed together with nine outgroup accessions. A phylogenetic tree was calculated using the maximum likelihood method. The tree showed that all groups that were found in previous studies were also present in our study. Additionally, seven new groups were identified based on the new haplotypes. Most B. tabaci haplotypes grouped based on their geographical origin. Two groups were found to have a worldwide distribution. Our results indicate that our knowledge on the species complex around B. tabaci is still far from complete.     

How different from random are docking predictions when ranked by scoring functions?

Docking algorithms predict the structure of protein–protein interactions. They sample the orientation of two unbound proteins to produce various predictions about their interactions, followed by a scoring step to rank the predictions. We present a statistical assessment of scoring functions used to rank near‐native orientations, applying our statistical analysis to a benchmark dataset of decoys of protein–protein complexes and assessing the statistical significance of the outcome in the Critical Assessment of PRedicted Interactions (CAPRI) scoring experiment. A P value was assigned that depended on the number of near‐native structures in the sampling. We studied the effect of filtering out redundant structures and tested the use of pair‐potentials derived using ZDock and ZRank. Our results show that for many targets, it is not possible to determine when a successful reranking performed by scoring functions results merely from random choice. This analysis reveals that changes should be made in the design of the CAPRI scoring experiment. We propose including the statistical assessment in this experiment either at the preprocessing or the evaluation step. Proteins 2010. © 2010 Wiley‐Liss, Inc.