Ligand Binding and Crystal Structures of the Substrate-Binding Domain of the ABC Transporter OpuA

Background

The ABC transporter OpuA from Lactococcus lactis transports glycine betaine upon activation by threshold values of ionic strength. In this study, the ligand binding characteristics of purified OpuA in a detergent-solubilized state and of its substrate-binding domain produced as soluble protein (OpuAC) was characterized.

Principal Findings

The binding of glycine betaine to purified OpuA and OpuAC (K_D = 4–6 µM) did not show any salt dependence or cooperative effects, in contrast to the transport activity. OpuAC is highly specific for glycine betaine and the related proline betaine. Other compatible solutes like proline and carnitine bound with affinities that were 3 to 4 orders of magnitude lower. The low affinity substrates were not noticeably transported by membrane-reconstituted OpuA. OpuAC was crystallized in an open (1.9 Å) and closed-liganded (2.3 Å) conformation. The binding pocket is formed by three tryptophans (Trp-prism) coordinating the quaternary ammonium group of glycine betaine in the closed-liganded structure. Even though the binding site of OpuAC is identical to that of its B. subtilis homolog, the affinity for glycine betaine is 4-fold higher.

Conclusions

Ionic strength did not affect substrate binding to OpuA, indicating that regulation of transport is not at the level of substrate binding, but rather at the level of translocation. The overlap between the crystal structures of OpuAC from L.lactis and B.subtilis, comprising the classical Trp-prism, show that the differences observed in the binding affinities originate from outside of the ligand binding site.     

WOX14 promotes bioactive gibberellin synthesis and vascular cell differentiation in Arabidopsis

Procambial and cambial stem cells provide the initial cells that allow the formation of vascular tissues. WOX4 and WOX14 have been shown to act redundantly to promote procambial cell proliferation and differentiation. Gibberellins (GAs), which have an important role in wood formation, also stimulate cambial cell division. Here we show that the loss of WOX14 function phenocopies some traits of GA‐deficient mutants that can be complemented by exogenous GA application, whereas WOX14 overexpression stimulates the expression of GA3ox anabolism genes and represses GA2ox catabolism genes, promoting the accumulation of bioactive GA. More importantly, our data clearly indicate that WOX14 but not WOX4 promotes vascular cell differentiation and lignification in inflorescence stems of Arabidopsis.     

Production of rhamnolipids in solid-state cultivation using a mixture of sugarcane bagasse and corn bran supplemented with glycerol and soybean oil

Rhamnolipid biosurfactants are attracting attention due to their low toxicity, high biodegradability, and good ecological acceptability. However, production in submerged culture is made difficult by severe foaming problems. Solid-state cultivation (SSC) is a promising alternative production method. In the current work, we report the optimization of rhamnolipid production by Pseudomonas aeruginosa UFPEDA 614 on a solid substrate containing sugarcane bagasse and corn bran. The best rhamnolipid production, 45 g/l of impregnating solution used, was obtained with a 50:50 (m/m) mixture of sugarcane bagasse and corn bran supplemented with an impregnating solution containing 6% (v/v) of each of glycerol and soybean oil. This level is comparable with those of previous studies undertaken in solid-state cultivation; the composition of the biosurfactant is similar, but our medium is cheaper. Our work therefore provides a suitable basis for future studies of the development of an SSC-based process for rhamnolipid production.     

The enlarged population of marginal zone/CD1d(high) B lymphocytes in nonobese diabetic mice maps to diabetes susceptibility region Idd11

The NOD mouse is an important experimental model for human type 1 diabetes. T cells are central to NOD pathogenesis, and their function in the autoimmune process of diabetes has been well studied. In contrast, although recognized as important players in disease induction, the role of B cells is not clearly understood. In this study we characterize different subpopulations of B cells and demonstrate that marginal zone (MZ) B cells are expanded 2- to 3-fold in NOD mice compared with nondiabetic C57BL/6 (B6) mice. The NOD MZ B cells displayed a normal surface marker profile and localized to the MZ region in the NOD spleen. Moreover, the MZ B cell population developed early during the ontogeny of NOD mice. By 3 wk of age, around the time when autoreactive T cells are first activated, a significant MZ B cell population of adult phenotype was found in NOD, but not B6, mice. Using an F2(B6 x NOD) cross in a genome-wide scan, we map the control of this trait to a region on chromosome 4 (logarithm of odds score, 4.4) which includes the Idd11 and Idd9 diabetes susceptibility loci, supporting the hypothesis that this B cell trait is related to the development of diabetes in the NOD mouse.     

Mutations in C2orf37, Encoding a Nucleolar Protein, Cause Hypogonadism, Alopecia, Diabetes Mellitus, Mental Retardation, and Extrapyramidal Syndrome

Hypogonadism, alopecia, diabetes mellitus, mental retardation, and extrapyramidal syndrome (also referenced as Woodhouse-Sakati syndrome) is a rare autosomal recessive multisystemic disorder. We have identified a founder mutation consisting of a single base-pair deletion in C2orf37 in eight families of Saudi origin. Three other loss-of-function mutations were subsequently discovered in patients of different ethnicities. The gene encodes a nucleolar protein of unknown function, and the cellular phenotype observed in patient lymphoblasts implicates a role for the nucleolus in the pathogenesis of this disease. Our findings expand the list of human disorders linked to the nucleolus and further highlight the developmental and/or maintenance functions of this organelle.     

Differential toxicity of TAR DNA‐binding protein 43 isoforms depends on their submitochondrial localization in neuronal cells

TAR DNA ‐binding protein 43 (TDP ‐43) is an RNA ‐binding protein and a major component of protein aggregates found in amyotrophic lateral sclerosis and several other neurodegenerative diseases. TDP ‐43 exists as a full‐length protein and as two shorter forms of 25 and 35 kD a. Full‐length mutant TDP ‐43s found in amyotrophic lateral sclerosis patients re‐localize from the nucleus to the cytoplasm and in part to mitochondria, where they exert a toxic role associated with neurodegeneration. However, induction of mitochondrial damage by TDP ‐43 fragments is yet to be clarified. In this work, we show that the mitochondrial 35 kD a truncated form of TDP ‐43 is restricted to the intermembrane space, while the full‐length forms also localize in the mitochondrial matrix in cultured neuronal NSC ‐34 cells. Interestingly, the full‐length forms clearly affect mitochondrial metabolism and morphology, possibly via their ability to inhibit the expression of Complex I subunits encoded by the mitochondrial‐transcribed mRNA s, while the 35 kD a form does not. In the light of the known differential contribution of the full‐length and short isoforms to generate toxic aggregates, we propose that the presence of full‐length TDP ‐43s in the matrix is a primary cause of mitochondrial damage. This in turn may cause oxidative stress inducing toxic oligomers formation, in which short TDP ‐43 forms play a major role.

     

Effect of meteorological parameters on Poaceae pollen in the atmosphere of Tetouan (NW Morocco)

Poaceae pollen is one of the most prevalent aeroallergens causing allergenic reactions. The aim of this study was to characterise the grass pollen season in Tetouan during the years 2008–2010, to analyse the effect of some meteorological parameters on the incidence of the airborne Poaceae pollen, and to establish forecasting variables for daily pollen concentrations. Aerobiological sampling was undertaken over three seasons using the volumetric method. The pollen season started in April and showed the highest pollen index in May and June, when the maximum temperature ranged from 23 to 27 °C, respectively. The annual pollen score recorded varied from year to year between 2,588 and 5,404. The main pollen season lasted 114–173 days, with peak days occurring mainly in May; the highest concentration reached 308 pollen grains/m³. Air temperature was the most important meteorological parameter and correlated positively to daily pollen concentration increase. An increase in relative humidity and precipitation was usually related to a decrease in airborne pollen content. External validation of the models performed using data from 2011 showed that Poaceae pollen concentration can be highly predicted (64.2–78.6 %) from the maximum temperature, its mean concentration for the same day in other years, and its concentration recorded on the previous day. Sensitive patients suffering allergy to Poaceae pollen are at moderate to highest risk of manifesting allergic symptoms to grass pollen over 33–42 days. The results obtained provide new information on the quantitative contribution of the Poaceae pollen to the airborne pollen of Tetouan and on its temporal distribution. Airborne pollen can be surveyed and forecast in order to warn the atopic population.     

Effect of adding plyometric training to physical education sessions on specific biomechanical parameters in primary school girls

Objectives:The study aimed to determine the effect of adding a school-based plyometric training program (PMT) to physical education (PE) sessions on the strength, balance, and flexibility in primary school girls.Methods:Students from grades 3-6 were randomized equally to a plyometric or control group. In the control group, students took their regular PE classes twice a week. In the plyometric group, students performed PMT twice a week during the initial 20 minutes of every PE session. The Lido Linea closed kinetic chain isokinetic dynamometer, Star excursion balance test (SEBT), and sit-and-reach test were used to assess muscle strength, balance, and flexibility, respectively, before and after nine weeks of training.Results:The improvement in extension peak force (p=0.04) and extension total work (p<0.001) was more prevalent in the PMT group than in the control group. SEBT scores had improved significantly (p<0.05) for all directions in the PMT group, except in the anterior direction, which was highly significant (p<0.001). Hamstring and lower back flexibility had improved more in the PMT group than in the control group (p<0.001).Conclusion:Adding PMT to regular PE classes has a positive and notable effect on muscle strength, balance, and flexibility in primary school students.