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Background Numerous reports have shown the role of human leukocyte antigen (HLA) alleles in the induction of cutaneous adverse drug reactions by moderating drug metabolism. We therefore aimed to investigate the docking patterns of four HLA alleles (HLA-B x 5101, HLA-B x 1501, HLA-A x 02:06 and HLA-B x 57:01) against four commercial drugs. Methodology Four drugs (phenytoin (PHT), amoxicillin (AMX), aceclofenac (ACE) and ciprofloxacin (CIP)) were investigated for their docking behavior against four HLA alleles (HLA-B x 5101, HLA-B x 1501, HLA-A x 02:06, and HLA-B x 57:01) using the SwissDock method. In addition, toxicity (Tox) and the search tool for interactions of chemicals (STITCH) (protein-drug interaction) analyses were also carried out using the predicating the small molecule pharmaco-kinetic (pk) properties using graph-based signature method (pkCSM) and STITCH free online servers, respectively. Results Toxicity analysis showed that two drugs (amoxicillin and ciprofloxacin) exhibit hepatotoxicity. The STITCH analysis of the drug amoxicillin revealed its interaction with two human proteins. The drug phenytoin exhibited the lowest binding energy (LBE) with all four HLA alleles (HLA-B x 5101, HLA-B x 1501, HLA-A x 02:06, and HLA-B x 57:01). Conclusions The present findings provide new knowledge about the four drugs (phenytoin (PHT), amoxicillin (AMX), aceclofenac (ACE) and ciprofloxacin (CIP)) and their binding affinities with HLA alleles, which may cause cutaneous adverse drug reactions.
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Background Sivelestat is a potent and specific neutrophil elastase inhibitor. It is clinically used in treating lung injury and respiratory distress syndrome. This engaged us to undertake the present study in which sivelestat was studied as an anti-inflammatory and anti-viral agent. Methodology The docking study of sivelestat on matrix metalloproteinase-2 (MMP-2), matrix metalloproteinase-9 (MMP-9), chikungunya virus nonstructural protein-2 (CVnsP2) protease, and influenza A (H1N9) virus neuraminidase was assessed using the Chemistry at Harvard Macromolecular Mechanics (CHARMM) Dock (CDOCK) method. Furthermore, molecular physicochemical; bioactivity; absorption, distribution, metabolism, and excretion (ADME); toxicity; and Search Tool for Interacting Chemicals (STITCH) analyses were performed by using the Molinspiration (Molinspiration Cheminformatics, Slovensky Grob, Slovak Republic), SwissADME SwissADME (Swiss Institute of Bioinformatics, Quartier Sorge - Bâtiment Amphipôle, Switzerland), pkCSM (University of Melbourne, Melbourne, Australia), and STITCH-free online tools. Results The molecular physicochemical assessment of the ligand (sivelestat) showed no (zero) violation and agreed with the thumb rule of five, otherwise known as Lipinski's rule of five. ADME prediction of the ligand (sivelestat) is shown to possess a low gastrointestinal absorption (GIA) property. Similarly, toxicity analysis of the ligand (sivelestat) is predicted to have a hepatotoxicity effect. STITCH analysis reveals that the ligand (sivelestat) has exhibited interactions with the three human proteins. Conclusions The present molecular docking studies showed that the ligand (sivelestat) has successfully docked with all four enzymes of interest. Hence, the current finding has provided a good understanding of sivelestat as an effective suppressor activity against all four enzymes: MMP-2, MMP-9, CVnsP2 protease, and influenza neuraminidase.
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Salt stress is one of the major factors impacting crop productivity worldwide. Through a variety of effector and signaling pathways, plants achieve survival under salinity stress by maintaining high cytosolic potassium/sodium ion (K+/Na+) ratios, preventing Na+ cytotoxicity, and retaining osmotic balance. Ras-related protein 5 (Rab5) members are involved in the trafficking of endosomes to the vacuole or plasma membrane (PM). The vacuolar protein sorting- associated protein 9 (vps9a) encodes the single guanine nucleotide exchange factor (GEF) that activates all three known Rab5 proteins in Arabidopsis thaliana. Previous work from our group has reported the critical function of vps9a for the operation of salt-induced endocytic pathway, as well as the expansion of endomembrane compartments under saline stress conditions. Here we show an additional role of vps9a in plant response to salt stress via maintenance of K+ status of the cell rather than Na+ homeostasis. Our results show that roots from vps9a-2 mutant, subjected to 100 mM NaCl, display alterations in transcript levels of genes involved in the K+ homeostasis pathway. Concurrent with the observed sensitivity of vps9a-2 mutant under NaCl stress, exposure to low K+ environments resulted in growth retardation, and reduced rate of endocytosis. Furthermore, vps9a-2 mutant displays reduced expression of auxin reporter, Direct Repeat-5 (DR5), and alterations in polarity and abundance of auxin efflux carrier PIN- FORMED2 (PIN2). Imposition of NaCl stress was found to be restrictive to the elongation capacity of cells in the root elongation zone of vps9a-2 mutant. Together our results indicate that alterations in K+ homeostasis and associated cellular changes causing increased cell wall pH, contribute to diminished root growth and compromised survival of vps9a-2 mutant under salt stress conditions.
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Plants challenged by limited phosphorus undergo dramatic morphological and architectural changes in their root systems in order to increase their absorptive surface area. In this paper, it is shown that phosphorus deficiency results in increased expression of the type I H+-pyrophosphatase AVP1 (AVP, Arabidopsis vacuolar pyrophosphatase), subsequent increased P-type adenosine triphosphatase (P-ATPase)-mediated rhizosphere acidification and root proliferation. Molecular genetic manipulation of AVP1 expression in Arabidopsis, tomato and rice results in plants that outperform controls when challenged with limited phosphorus. However, AVP1 over-expression and the resulting rhizosphere acidification do not result in increased sensitivity to AlPO4, apparently because of the enhancement of potassium uptake and the release of organic acids. Thus, the over-expression of type I H+-pyrophosphatases appears to be a generally applicable technology to help alleviate agricultural losses in low-phosphorus tropical/subtropical soils and to reduce phosphorus runoff pollution of aquatic and marine environments resulting from fertilizer application.
Assuntos
Arabidopsis/enzimologia , Pirofosfatase Inorgânica/metabolismo , Oryza/enzimologia , Fosfatos/metabolismo , Bombas de Próton/metabolismo , Solanum lycopersicum/enzimologia , Adaptação Fisiológica , Arabidopsis/genética , Arabidopsis/fisiologia , Proteínas de Arabidopsis , Biomassa , Ácidos Carboxílicos/metabolismo , Expressão Gênica , Concentração de Íons de Hidrogênio , Pirofosfatase Inorgânica/genética , Solanum lycopersicum/genética , Solanum lycopersicum/fisiologia , Oryza/genética , Oryza/fisiologia , Raízes de Plantas/crescimento & desenvolvimento , Raízes de Plantas/metabolismo , Brotos de Planta/crescimento & desenvolvimento , Plantas Geneticamente Modificadas/enzimologia , Plantas Geneticamente Modificadas/crescimento & desenvolvimento , Plantas Geneticamente Modificadas/metabolismo , Potássio/metabolismo , Bombas de Próton/genética , ATPases Translocadoras de Prótons/metabolismo , Sementes/crescimento & desenvolvimentoRESUMO
Salt stress is an environmental factor that severely impairs plant growth and productivity. We have cloned a novel isoform of a vacuolar Na+/H+ antiporter from Pennisetum glaucum (PgNHX1) that contains 5 transmembrane domains in contrast to AtNHX1 and OsNHX1 which have 9 transmembrane domains. Recently we have shown that PgNHX1 could confer high level of salinity tolerance when overexpressed in Brassica juncea. Here,we report the functional validation of this antiporter in crop plant rice. Overexpression of PgNHX1 conferred high level of salinity tolerance in rice. Transgenic rice plants overexpressing PgNHX1 developed more extensive root system and completed their life cycle by setting flowers and seeds in the presence of 150 mM NaCl. Our data demonstrate the potential of PgNHX1 for imparting enhanced salt tolerance capabilities to salt-sensitive crop plants for growing in high saline areas.
Assuntos
Oryza/efeitos dos fármacos , Oryza/metabolismo , Pennisetum/genética , Cloreto de Sódio/farmacologia , Trocadores de Sódio-Hidrogênio/genética , Trocadores de Sódio-Hidrogênio/metabolismo , Clonagem Molecular , Relação Dose-Resposta a Droga , Regulação da Expressão Gênica de Plantas , Germinação , Oryza/genética , Oryza/crescimento & desenvolvimento , Folhas de Planta/efeitos dos fármacos , Folhas de Planta/metabolismo , Plantas Geneticamente Modificadas , Isoformas de Proteínas/química , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Reprodutibilidade dos Testes , Trocadores de Sódio-Hidrogênio/químicaRESUMO
We evaluated DNA binding of the B-HLH family members TCF4 and USF1 using protein binding microarrays (PBMs) containing double-stranded DNA probes with cytosine on both strands or 5-methylcytosine (5mC) or 5-hydroxymethylcytosine (5hmC) on one DNA strand and cytosine on the second strand. TCF4 preferentially bound the E-box motif (CAN|NTG) with strongest binding to the 8-mer CAG|GTGGT. 5mC uniformly decreases DNA binding of both TCF4 and USF1. The bulkier 5hmC also inhibited USF1 binding to DNA. In contrast, 5hmC dramatically enhanced TCF4 binding to E-box motifs ACAT|GTG and ACAC|GTG, being better bound than any 8-mer containing cytosine. Examination of X-ray structures of the closely related TCF3 and USF1 bound to DNA suggests TCF3 can undergo a conformational shift to preferentially bind to 5hmC while the USF1 basic region is bulkier and rigid precluding a conformation shift to bind 5hmC. These results greatly expand the regulatory DNA sequence landscape bound by TCF4.
Assuntos
5-Metilcitosina/análogos & derivados , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/química , DNA/química , Elementos E-Box , Análise Serial de Proteínas/métodos , Fator de Transcrição 4/química , 5-Metilcitosina/química , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos/ultraestrutura , Sítios de Ligação , DNA/ultraestrutura , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/ultraestrutura , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Mapeamento de Interação de Proteínas , Fator de Transcrição 4/ultraestruturaRESUMO
We have cloned and characterized the cDNA, genomic clone and upstream promoter region of a vacuolar ATPase (V-ATPase) c subunit (PgVHA-c1) from Pennisetum glaucum. The deduced amino acid sequence shows 98-71% sequence identity with V-ATPase from rice and Arabidopsis, and is a highly hydrophobic protein with four transmembrane regions. PgVHA-c1-GFP fusion protein is expressed in BY2 cells on the endo-membranes surrounding vacuoles; however, PgVHA-c1 could not functionally complement V-ATPase-c deletion mutants of yeast. The sequence analysis of the genomic clone revealed the presence of two introns in the coding region, and the splice junctions followed the typical canonical GU-AG consensus sequence. The transcript analysis showed that the expression of PgVHA-c1 was stimulated more in response to salinity stress and very marginally in response to drought and low temperature stress. Exogenous application of abscisic acid, salicylic acid and calcium stimulated the transcript level in the absence of stress. We have cloned the 5'-flanking regions of PgVHA-c1 and mapped its transcript start site at 78 bp upstream of ATG. Transgenic tobacco with promoter::GUS constructs showed that the region -288/+78 was sufficient for GUS expression. The expression of the reporter gene even with the full-length promoter was limited to shoot hairs and to male and female reproductive organs. The dehydration-responsive element (DRE) and ABA-responsive element (ABRE) in the promoter did not show consensus flanking regions; however, gel mobility shift assays showed that Pennisetum has specific transacting factors that showed binding to the core DRE, ABRE and TCA elements.