A theoretical analysis of the binding of palmitate by human serum albumin

The distribution of palmitate between the form bound by human serum albumin and the free form in plasma was calculated by use of 12 stepwise equilibrium constants and a computer program. Computations were carried out for molar ratios of palmitate to serum albumin of 0.5, 1,2,3, and 4. At most 0.0003% of the palmitate would be in the unbound form, and the remainder distributed among different complexes with albumin. At low molar ratios, the complexes with 1 to 2 moles of palmitate/albumin would predominate while at the highest ratio the complexes of 3 and 4 moles of palmitate/albumin would be most abundant. In the delivery of palmitate to tissues the relative contribution of the different complexes would change, as the molar ratio of fatty acid to albumin changed.     

Single-molecule force spectroscopy: optical tweezers, magnetic tweezers and atomic force microscopy

Single-molecule force spectroscopy has emerged as a powerful tool to investigate the forces and motions associated with biological molecules and enzymatic activity. The most common force spectroscopy techniques are optical tweezers, magnetic tweezers and atomic force microscopy. Here we describe these techniques and illustrate them with examples highlighting current capabilities and limitations.     

Insulation and wiring specificity of BceR‐like response regulators and their target promoters in Bacillus subtilis

BceRS and PsdRS are paralogous two‐component systems in Bacillus subtilis controlling the response to antimicrobial peptides. In the presence of extracellular bacitracin and nisin, respectively, the two response regulators (RRs) bind their target promoters, P_bceA or P_psdA, resulting in a strong up‐regulation of target gene expression and ultimately antibiotic resistance. Despite high sequence similarity between the RRs BceR and PsdR and their known binding sites, no cross‐regulation has been observed between them. We therefore investigated the specificity determinants of P_bceA and P_psdA that ensure the insulation of these two paralogous pathways at the RR–promoter interface. In vivo and in vitro analyses demonstrate that the regulatory regions within these two promoters contain three important elements: in addition to the known (main) binding site, we identified a linker region and a secondary binding site that are crucial for functionality. Initial binding to the high‐affinity, low‐specificity main binding site is a prerequisite for the subsequent highly specific binding of a second RR dimer to the low‐affinity secondary binding site. In addition to this hierarchical cooperative binding, discrimination requires a competition of the two RRs for their respective binding site mediated by only slight differences in binding affinities.     

Turkish graveyards as refuges for orchids against tuber harvest

Harvest of orchid tubers for salep production is widespread in southwestern Asia and the Balkans and constitutes a major conservation risk for wild orchid populations. Synanthropic habitats, such as graveyards, are important refuges for orchids and other organisms and could offer protection from salep harvesting because of their special cultural role. However, little is known about the occurrence and factors influencing harvesting of salep in graveyards. During field surveys of 474 graveyards throughout Turkey, we observed 333 graveyards with orchids, 311 graveyards with tuberous orchids, and salep harvest in 14 graveyards. Altogether, 530 individuals of 17 orchid species were collected, representing 9% of the individuals recorded. Harvesting intensity was relatively low, and populations were usually not wholly destroyed. However, some species were clearly more affected than others. Salep harvesting risk of orchid species was significantly associated with flowering time, with early‐flowering species being more affected. A marginally significant positive relationship between harvesting risk and species‐specific tuber size was also detected. Our data suggest that graveyards might offer some protection against salep harvesting in Turkey, but they also show that some orchid taxa are much more affected than others. Overall, our observations add more weight to the conservation value of these special habitats.     

An Improved Sequence-aided T-RFLP Analysis of Bacterial Succession During Oyster Mushroom Substrate Preparation

While oyster mushroom (Pleurotus spp.) is one of the most popular cultivated edible mushrooms, there is scanty information about the microbial community taking part in mushroom substrate production. In this study, an improved sequence-aided terminal restriction fragment length polymorphism (T-RFLP) was used to identify and (semi-)quantify the dominant bacteria of oyster mushroom substrate preparation. The main features of the improved T-RFLP data analysis were the alignment of chromatograms with variable clustering thresholds, the visualization of data matrix with principal component analysis ordination superimposed with cluster analysis, and the search for stage-specific peaks (bacterial taxa) with similarity percentage (analysis of similarity) analysis, followed by identification with clone libraries. By applying this method, the dominance of the following bacterial genera was revealed during oyster mushroom substrate preparation: Pseudomonas and Sphingomonas at startup, Bacillus, Geobacillus, Ureibacillus, Pseudoxanthomonas, and Thermobispora at the end of partial composting, and finally several genera of Actinobacteria, Thermus, Bacillus, Geobacillus, Thermobacillus, and Ureibacillus in the mature substrate. As the proportion of uncultured bacteria increased during the process, it is worth establishing strain collections from partial composting and from mature substrate for searching new species.     

Geographic differences in genetic susceptibility to IgA nephropathy: GWAS replication study and geospatial risk analysis

IgA nephropathy (IgAN), major cause of kidney failure worldwide, is common in Asians, moderately prevalent in Europeans, and rare in Africans. It is not known if these differences represent variation in genes, environment, or ascertainment. In a recent GWAS, we localized five IgAN susceptibility loci on Chr.6p21 (HLA-DQB1/DRB1, PSMB9/TAP1, and DPA1/DPB2 loci), Chr.1q32 (CFHR3/R1 locus), and Chr.22q12 (HORMAD2 locus). These IgAN loci are associated with risk of other immune-mediated disorders such as type I diabetes, multiple sclerosis, or inflammatory bowel disease. We tested association of these loci in eight new independent cohorts of Asian, European, and African-American ancestry (N = 4,789), followed by meta-analysis with risk-score modeling in 12 cohorts (N = 10,755) and geospatial analysis in 85 world populations. Four susceptibility loci robustly replicated and all five loci were genome-wide significant in the combined cohort (P = 5×10⁻³²–3×10⁻¹⁰), with heterogeneity detected only at the PSMB9/TAP1 locus (I² = 0.60). Conditional analyses identified two new independent risk alleles within the HLA-DQB1/DRB1 locus, defining multiple risk and protective haplotypes within this interval. We also detected a significant genetic interaction, whereby the odds ratio for the HORMAD2 protective allele was reversed in homozygotes for a CFHR3/R1 deletion (P = 2.5×10⁻⁴). A seven–SNP genetic risk score, which explained 4.7% of overall IgAN risk, increased sharply with Eastward and Northward distance from Africa (r = 0.30, P = 3×10⁻¹²⁸). This model paralleled the known East–West gradient in disease risk. Moreover, the prediction of a South–North axis was confirmed by registry data showing that the prevalence of IgAN–attributable kidney failure is increased in Northern Europe, similar to multiple sclerosis and type I diabetes. Variation at IgAN susceptibility loci correlates with differences in disease prevalence among world populations. These findings inform genetic, biological, and epidemiological investigations of IgAN and permit cross-comparison with other complex traits that share genetic risk loci and geographic patterns with IgAN.     

Synchronous Elicitation of Development in Root Caps Induces Transient Gene Expression Changes Common to Legume and Gymnosperm Species

Root cap development in cereals and legumes is self-regulated by a repressor that accumulates in the extracellular environment, and immersing the root tip into water results in renewed cap development. By exploiting this phenomenon, root cap mitosis and differentiation can be synchronously induced among populations. In Pisum sativum L., messenger RNA (mRNA) differential display revealed changes in expression of approximately 1% of the sample mRNA population within minutes of induced cap turnover. This profile changes sequentially over a period of 30 min, then stabilizes. Microarray analysis of Medicago truncatula root caps confirmed changes in expression of approximately 1% of the target population, within minutes. A cell specific marker for cap turnover exhibited the same temporal and spatial expression profile in the gymnosperm species Norway spruce (Picea abies) as in pea. Induced cap development provides a means to profile cell-specific gene expression among phylogenetically diverse species from the early moments of mitosis and cellular differentiation.