Structural insight into the dual ligand specificity and mode of high density lipoprotein association of apolipoprotein D

Human apolipoprotein D (ApoD) occurs in plasma associated with high density lipoprotein. Apart from the involvement in lipid metabolism, its binding activity for progesterone and arachidonic acid plays a role in cancer development and neurological diseases. The crystal structures of free ApoD and its complex with progesterone were determined at 1.8A resolution and reveal a lipocalin fold. The narrow, mainly uncharged pocket within the typical beta-barrel accommodates progesterone with its acetyl side chain oriented toward the bottom. The cavity adopts essentially the same shape in the absence of progesterone and allows complexation of arachidonic acid as another cognate ligand. Three of the four extended loops at the open end of the beta-barrel expose hydrophobic side chains, which is an unusual feature for lipocalins and probably effects association with the high density lipoprotein particle by mediating insertion into the lipid phase. This mechanism is in line with an unpaired Cys residue in the same surface region that can form a disulfide cross-link with apolipoprotein A-II.     

Nucleocytoplasmic shuttling of the zinc finger protein EZI Is mediated by importin-7-dependent nuclear import and CRM1-independent export mechanisms

Nucleocytoplasmic translocation constitutes a foundation for nuclear proteins to exert their proper functions and hence for various biological reactions to occur normally in eukaryotic cells. We reported previously that EZI/Zfp467, a 12 zinc finger motif-containing protein, localizes predominantly in the nucleus, yet the underlying mechanism still remains elusive. Here we constructed a series of mutant forms of EZI and examined their subcellular localization. The results delineated a non-canonical nuclear localization signal in the region covering the 9th to the 12th zinc fingers, which was necessary for nuclear accumulation of EZI as well as sufficient to confer nuclear localizing ability to a heterologous protein. We also found that the N-terminal domain of EZI is necessary for its nuclear export, the process of which was not sensitive to the CRM1 inhibitor leptomycin B. An interaction proteomics approach and the following co-immunoprecipitation experiments identified the nuclear import receptor importin-7 as a molecule that associated with EZI and, importantly, short interfering RNA-mediated knockdown of importin-7 expression completely abrogated nuclear accumulation of EZI. Taken together, these results identify EZI as a novel cargo protein for importin-7 and demonstrate a nucleocytoplasmic shuttling mechanism that is mediated by importin-7-dependent nuclear localization and CRM1-independent nuclear export.     

Effects of fermented oyster mushroom ( Pleurotus ostreats) by-product supplementation on growth performance, blood parameters and meat quality in finishing Berkshire pigs

The objective of the present study was to investigate the effects of fermented oyster mushroom (Pleurotus ostreats) by-production (FOMP) supplementation on the growth performance, blood parameters, carcass traits and meat quality in finishing Berkshire pigs. FOMP was made by mixing oyster mushroom by-production with rice bran and barley bran and this mixture was fermented for 60 days. The experimental diets were 0, 3, 5 and 7% of FOMP added to C, T1, T2 and T3 in the basis diet for 7 weeks. Average daily gain (kg/day) was higher in C and T1 than in T2 and T3 ( P < 0.05). Average daily feed intake (kg/day) and feed conversion increased by the addition of FOMP ( P < 0.05). Total cholesterol and high-density lipoprotein cholesterol were higher in T3 than other treatments ( P < 0.05). Carcass weight (kg) was higher in C and T1 than in T2 and T3 ( P < 0.05). Dressing (%) was higher in C than in T3 ( P < 0.05). Crude protein was lower in T3 than in other treatments ( P < 0.05). Crude fat was higher in T2 and T3 than in C ( P < 0.05). pH₂₄ was higher in C than in other treatments ( P < 0.05). Cooking loss (%) was higher in T1 than T2 ( P < 0.05). Water-holding capacity (%) was higher in C than in T1 ( P < 0.05). In meat colour, CIE a* was lower by the addition of FOMP ( P < 0.05). CIE b* was higher in C than in other treatments ( P < 0.05). In backfat colour, CIE L* was lower in T3 than other treatments ( P < 0.05). CIE b* was lower by addition of FOMP ( P < 0.05). Palmitoleic and oleic acid were higher in T3 than in other treatments ( P < 0.05). Linoleic and arachidonic acids were higher in T2 than in other treatments ( P < 0.05). The results indicate that 3% of FOMP affected the growth performance, carcass traits, meat quality and fatty acid in contrast to addition of 5% of FOMP for Berkshire pigs during the finishing period.     

High frequency somatic embryogenesis and plant regeneration in root explant cultures of carnation

Root explants excised from carnation plants maintained in vitro formed off-white, friable calluses after three weeks of culture on Murashige and Skoog (MS) medium supplemented with 1 mg l⁻¹ thidiazuron (TDZ) and 1 mg l⁻¹ α-naphthalaneacetic acid (NAA). These calluses were subsequently transferred to MS basal medium where, after an additional four weeks of culture, approximately 50% of the calluses formed somatic embryos. However, calluses formed on root explants that had been cultured on MS medium supplemented with 2,4-dichlorophenoxyacetic acid did not produce somatic embryos upon transfer to MS basal medium. Somatic embryos developed into plantlets and subsequently were grown to maturity. These results indicate that root explants have a high competence for somatic embryogenesis in carnation. J. Seo and S.W. Kim contributed equally to this work.     

Diagnosing idiopathic learning disability: a cost-effectiveness analysis of microarray technology in the National Health Service of the United Kingdom

Array based comparative genomic hybridisation (aCGH) is a powerful technique for detecting clinically relevant genome imbalance and can offer 40 to > 1000 times the resolution of karyotyping. Indeed, idiopathic learning disability (ILD) studies suggest that a genome-wide aCGH approach makes 10–15% more diagnoses involving genome imbalance than karyotyping. Despite this, aCGH has yet to be implemented as a routine NHS service. One significant obstacle is the perception that the technology is prohibitively expensive for most standard NHS clinical cytogenetics laboratories. To address this, we investigated the cost-effectiveness of aCGH versus standard cytogenetic analysis for diagnosing idiopathic learning disability (ILD) in the NHS. Cost data from four participating genetics centres were collected and analysed. In a single test comparison, the average cost of aCGH was ￡442 and the average cost of karyotyping was ￡117 with array costs contributing most to the cost difference. This difference was not a key barrier when the context of follow up diagnostic tests was considered. Indeed, in a hypothetical cohort of 100 ILD children, aCGH was found to cost less per diagnosis (￡3,118) than a karyotyping and multi-telomere FISH approach (￡4,957). We conclude that testing for genomic imbalances in ILD using microarray technology is likely to be cost-effective because long-term savings can be made regardless of a positive (diagnosis) or negative result. Earlier diagnoses save costs of additional diagnostic tests. Negative results are cost-effective in minimising follow-up test choice. The use of aCGH in routine clinical practice warrants serious consideration by healthcare providers. Copyright statement The Corresponding Author has the right to grant on behalf of all authors and does grant on behalf of all authors, an exclusive licence (or non exclusive for government employees) on a worldwide basis to the BMJ Publishing Group Ltd, and its Licensees to permit this article (if accepted) to be published in BMJ editions and any other BMJPGL products and to exploit all subsidiary rights, as set out in our licence (bmj.com/advice/copyright.shtml). Authorship The authors included on this paper fulfil the criteria of authorship and no one who fulfils the criteria has been excluded from authorship. The authors made a substantial contribution to the conception, design, analysis and interpretation of data. They were involved in drafting the article or revising it critically for important intellectual content and approving the version to be published. Contributorship Sarah Wordsworth (Guarantor): Planning, conducting and reporting work, interpretation of data, drafting and revising article. James Buchanan: Conducting and reporting work, interpretation of data, revising article. Regina Regan: Completing costing questionnaire, providing protocol details, other costing information, interpretation of data, information about learning disability and genome imbalance and revising article. Val Davison: Completing costing questionnaire, providing protocol details, sharing overall laboratory experience and drafting article. Kim Smith: Completing costing questionnaire, providing protocol details, drafting article. Sara Dyer: Completing costing questionnaire and providing protocol details. Carolyn Campbell: Completing costing questionnaire and providing protocol details. Edward Blair: Critical appraisal of article for clinical content and revising article. Eddy Maher: Completing costing questionnaire, providing protocol details, sharing overall laboratory experience and drafting article. Jenny Taylor: Planning and facilitating work between centres. Drafting and revising article. Samantha JL Knight: Completing costing questionnaire, providing protocol details, other costing information, interpretation of data, providing information about learning disability and genome imbalance, drafting and revising article. Jenny Taylor and Samantha JL Knight contributed equally to the work presented.     

Improved insulin sensitivity and adiponectin level after exercise training in obese Korean youth

Objective: The objective of this study was to investigate the association among adiposity, insulin resistance, and inflammatory markers [high‐sensitivity C‐reactive protein (hs‐CRP), interleukin (IL)‐6, and tumor necrosis factor (TNF)‐α] and adiponectin and to study the effects of exercise training on adiposity, insulin resistance, and inflammatory markers among obese male Korean adolescents. Research Methods and Procedures: Twenty‐six obese and 14 lean age‐matched male adolescents were studied. We divided the obese subjects into two groups: obese exercise group (N = 14) and obese control group (N = 12). The obese exercise group underwent 6 weeks of jump rope exercise training (40 min/d, 5 d/wk). Adiposity, insulin resistance, lipid profile, hs‐CRP, IL‐6, TNF‐α, and adiponectin were measured before and after the completion of exercise training. Results: The current study demonstrated higher insulin resistance, total cholesterol, LDL‐C levels, triglyceride, and inflammatory markers and lower adiponectin and HDL‐C in obese Korean male adolescents. Six weeks of increased physical activity improved body composition, insulin sensitivity, and adiponectin levels in obese Korean male adolescents without changes in TNF‐α, IL‐6, and hs‐CRP. Discussion: Obese Korean male adolescents showed reduced adiponectin levels and increased inflammatory cytokines. Six weeks of jump rope exercise improved triglyceride and insulin sensitivity and increased adiponectin levels.     

Effects of pH on structural and functional properties of porin from the outer membrane of <Emphasis Type="Italic">Yersinia pseudotuberculosis</Emphasis>. I. Functionally important conformational transitions of yersinin

The changes in the structural and functional properties of yersinin, a porin from the outer membrane of Yersinia pseudotuberculosis, were studied in the pH range 8.0–2.0 using SDs-PAGE, scanning microcalorimetry, optical spectroscopy and bilayer lipid membrane technique. It was found that in the pH range under study the changes in the spatial structure of yersinin were biphasic. In the first steps of pH titration (pH 8.0–4.5), porin underwent a series of conformational transitions, which did not affect the trimeric structure of its molecule. In the second step (pH 4.0–2.0), structural rearrangements led to dissociation of the protein trimers into monomers. It is noteworthy that complete unfolding of the polypeptide chain of the protein was not observed even at low values of pH. Thus, at pH 2.0 the conformational intermediate of the protein retained up to 50% of its regular secondary structure. Studies of current fluctuations in the bilayer lipid membrane revealed that in weakly acidic media the conductivity of yersinin pores was decreased by one order of magnitude. The most drastic changes in the conductivity of the model membrane were observed at pH 5.8, whereas a further decrease of pH to 5.0 resulted in the closure of porin channels. It was concluded that the observed changes in the pore-forming properties of yersinin in a narrow range of pH represent an early step in the adaptation of bacteria to the changing conditions of the environment and entail control over the biosynthesis of nonspecific porins. The pH-dependent changes in the structure and pore-forming properties of yersinin provide additional evidence in favor of conformational and functional plasticity of porins.     

Screening of rifamycin producing marine sponge bacteria by LC-MS-MS

HPLC-MS-MS has been used for the identification and characterisation of rifamycin B and rifamycin SV in various strains of the marine sponge-derived bacterium Salinispora. Gradient elution using acetonitrile/water/ammonium acetate was used to separate the rifamycins from the matrix and negative ion-electrospray mass spectrometry was used for detection and confirmation. The presence of rifamycin in bacterial extracts was confirmed by matching retention times, parent ion spectra and the fragmentated parent ion spectra of the standard compounds and the bacterial extracts. All strains of the marine sponge bacterium Salinispora tested were found to contain rifamycin thus an alternate actinobacterial source of rifamycin was established.