Glyoxylate metabolism and fatty acid oxidation in mango fruit during development and ripening

Throughout the development (maturation) of mango fruit the contents of citric and glyoxylic acids increased steadily. As the fruit matured the levels of isocitrate lyase, malate lyase and alanine: glyoxylate aminotransferase increased and reached maximum values prior to the time of harvesting. At and after harvest the levels of malate lyase and alanine : glyoxylate aminotransferase began to decrease but that of isocitrate lyase remained high until after the harvest when it decreased. The level of glyoxylate reductase was highest in the early developmental stage but declined as the fruit matured and ripened. As the fruit ripened, after harvest, the amounts of citric and glyoxylic acids decreased concomitant with a considerable increase in the levels of isocitrate dehydrogenase, malic dehydrogenase, malic enzyme and glyoxylate dehydrogenase.Fatty acid oxidizing capacity of mitochondria isolated from immature (developing) and postclimacteric fruit pulps was much less than that observed with mitochondria from preclimacteric and climacteric fruit. Glyoxylate stimulated the oxidation of caprylic, lauric, myristic and palmitic acids and inhibited the activity of isocitrate dehydrogenase in vitro.     

Interaction of aromatic donor molecules with manganese(III) reconstituted horseradish peroxidase: proton nuclear magnetic resonance and optical difference spectroscopic studies

The interaction of aromatic donor molecules with manganese(III) protoporphyrin-apohorseradish peroxidase complex [Mn(III)HRP] was investigated by optical difference spectroscopy and relaxation rate measurements of 1H resonances of aromatic donor molecules (at 500 MHz). pH dependence of substrate proton resonance line-widths indicated that the binding was facilitated by protonation of an amino acid residue (with a pKa of 6.1), which is presumably distal histidine. Dissociation constants were evaluated from both optical difference spectroscopy and 1H-NMR relaxation measurements (pH 6.1). The dissociation constants of aromatic donor molecules were not affected by the presence of excess of I-, CN- and SCN-. From competitive binding studies it was shown that all these aromatic donor molecules bind to Mn(III)HRP at the same site, which is different from the binding site of I-, CN- and SCN-. Comparison of the dissociation constants between the different substrates suggests that hydrogen bonding of the donors with distal histidyl amino acid and hydrophobic interaction between the donors and active site contribute significantly towards the associating forces. Free energy, entropy and enthalpy changes associated with the Mn(III)HRP-substrate equilibrium have been evaluated. These thermodynamic parameters were found to be all negative. Distances of the substrate protons from the paramagnetic manganese ion of Mn(III)HRP were found to be in the range of 7.7 to 9.4 A. The Kd values, the thermodynamic parameters and the distances of the bound aromatic donor protons from metal center in the case of Mn(III)HRP were found to be very similar as in the case of native Fe(III)HRP.     

From LC-MS/MS metabolomics profiling of Kanchanara Guggulu to molecular docking and dynamics simulation of quercetin pentaacetate with aldose reductase

Kanchanara Guggulu (KG) is an important traditional medicine that is prescribed by the Ayurveda physicians for the treatment of swellings in various organs such as the thyroid, and lymph nodes. High-resolution mass-spectrometry-based metabolomics found metabolites in KG. LC-MS/MS-based metabolomics analysis of KG identified 2,579 compounds including quercetin and kaempferol derivatives. The molecular docking and dynamics analysis of quercetin pentaacetate with aldose reductase is documented for further consideration in drug discovery.     

A family of ubiquitin-like proteins binds the ATPase domain of Hsp70-like Stch

We have isolated two human ubiquitin-like (UbL) proteins that bind to a short peptide within the ATPase domain of the Hsp70-like Stch protein. Chap1 is a duplicated homologue of the yeast Dsk2 gene that is required for transit through the G2/M phase of the cell cycle and expression of the human full-length cDNA restored viability and suppressed the G2/M arrest phenotype of dsk2Delta rad23Delta Saccharomyces cerevisiae mutants. Chap2 is a homologue for Xenopus scythe which is an essential component of reaper-induced apoptosis in egg extracts. While the N-terminal UbL domains were not essential for Stch binding, Chap1/Dsk2 contains a Sti1-like repeat sequence that is required for binding to Stch and is also conserved in the Hsp70 binding proteins, Hip and p60/Sti1/Hop. These findings extend the association between Hsp70 members and genes encoding UbL sequences and suggest a broader role for the Hsp70-like ATPase family in regulating cell cycle and cell death events.     

Sensitivity of nisin-resistant Listeria monocytogenes to heat and the synergistic action of heat and nisin

Nisin, a bacteriocin produced by some strains of Lactococcus lactis, acts against foodborne pathogen Listeria monocytogenes. A single exposure of cells to nisin can generate nisin-resistant (Nisr) mutants, which may compromise the use of nisin in the food industry. The objective of this research was to compare the heat resistance of Nisr and wild type (WT) Listeria monocytogenes. The synergistic effect of heat-treatment (55 degrees C) and nisin (500 IU ml-1) on the Nisr cells and the WT L. monocytogenes Scott A was also studied. When the cells were grown in the absence of nisin, there was no significant (alpha = 0.05) difference in heat resistance between WT and Nisr cells of L. monocytogenes at 55, 60 and 65 degrees C. However, when the Nisr cells were grown in the presence of nisin, they were more sensitive to heat at 55 degrees C than the WT cells. The D-values at 55 degrees C were 2.88 and 2.77 min for Nisr ATCC 700301 and ATCC 700302, respectively, which was significantly (alpha = 0.05) lower than the D-value for WT, 3.72 min. When Nisr cells were subjected to a combined treatment of heat and nisin, there was approximately a four log reduction during the first 7 min of treatment.     

Genetic mapping of leucine and isoleucine-valine loci in Rhizobium japonicum.

Leucine and isoleucine-valine loci have been mapped in Rhizobium japonicum. Transformation analysis suggests a common pathway for isoleucine-valine biosynthesis. Three-point reciprocal crosses indicated that all the leucine loci are not genetically linked.     

Remission of Collagen-Induced Arthritis through Combination Therapy of Microfracture and Transplantation of Thermogel-Encapsulated Bone Marrow Mesenchymal Stem Cells

The persistent inflammation of rheumatoid arthritis (RA) always leads to partial synovial hyperplasia and the destruction of articular cartilage. Bone marrow mesenchymal stem cells (BMMSCs) have been proven to possess immunosuppressive effects, and widely explored in the treatment of autoimmune diseases. However, poor inhibitory effect on local inflammatory state and limited capacity of preventing destruction of articular cartilage by systemic BMMSCs transplantation were observed. Herein, toward the classical type II collagen-induced arthritis in rats, the combination treatment of microfracture and in situ transplantation of thermogel-encapsulated BMMSCs was verified to obviously down-regulate the ratio of CD4⁺ to CD8⁺ T lymphocytes in peripheral blood. In addition, it resulted in the decreased levels of inflammatory cytokines, such as interleukin-1β, tumor necrosis factor-α and anti-collagen type II antibody, in the serum. Simultaneously, the combination therapy also could inhibit the proliferation of antigen specific lymphocytes and local joint inflammatory condition, and prevent the articular cartilage damage. The results indicated that the treatment programs could effectively stimulate the endogenous and exogenous BMMSCs to exhibit the immunosuppression and cartilage protection capability. This study provided a new therapeutic strategy for autoimmune inflammatory diseases, such as RA.     

Discovery of a Natural Product-Like iNOS Inhibitor by Molecular Docking with Potential Neuroprotective Effects In Vivo

In this study, we applied structure-based virtual screening techniques to identify natural product or natural product-like inhibitors of iNOS. The iNOS inhibitory activity of the hit compounds was characterized using cellular assays and an in vivo zebrafish larvae model. The natural product-like compound 1 inhibited NO production in LPS-stimulated Raw264.7 macrophages, without exerting cytotoxic effects on the cells. Significantly, compound 1 was able to reverse MPTP-induced locomotion deficiency and neurotoxicity in an in vivo zebrafish larval model. Hence, compound 1 could be considered as a scaffold for the further development of iNOS inhibitors for potential anti-inflammatory or anti-neurodegenerative applications.     

Structural analysis and molecular dynamics simulations of novel δ-endotoxin Cry1Id from Bacillus thuringiensis to pave the way for development of novel fusion proteins against insect pests of crops

The theoretical three-dimensional structure of a novel δ-endotoxin Cry1Id (81 kDa) belonging to Cry1I class, toxic to many of the lepidopteran pests has been investigated through comparative modeling. Molecular dynamics (MD) simulations was carried out to characterize its structural and dynamical features at 10 ns in explicit solvent using the GROMACS version 4.5.4. Finally the simulated model was validated by the SAVES, WHAT IF, MetaMQAP, ProQ, ModFOLD and MolProbity servers. Despite low sequence identity with its structural homologs, Cry1Id not only resembles the previously reported Cry structures but also shares the common five conserved blocks of amino acid residues. Although the domain II of Cry1Id superpose well with its closest structural homolog Cry8Ea1, variation of amino acids and length in the apical loop2 of domain II was observed. In this work, we have hypothesized that the variations in apical loop2 might be the sole factor for providing variable surface accessibility to Cry1Id protein that could be important in receptor recognition. MD simulation showed the proposed endotoxin retains its stable conformation in aqueous solution. The result from this study is expected to aid in the development hybrid Cry proteins and new potent fusion proteins with novel specificities against different insect pests for improved pest management of crop plants.