L-Arginine and tetrahydrobiopterin supported nitric oxide production is crucial for the microbicidal activity of neutrophils

Recent report from this lab has shown role of Rac2 in the translocation of inducible nitric oxide synthase (iNOS) to the phagosomal compartment of polymorphonuclear leukocytes (PMNs) following phagocytosis of beads. This study was undertaken to further assess the status and role of tetrahydrobiopterin (BH₄), a redox-sensitive cofactor, L-arginine, and the substrate of nitric oxide synthase (NOS) in sustained nitric oxide (˙NO) production in killing of phagocytosed microbes (Escherichia coli) by human PMNs. Time-dependent study revealed consistent NO and reactive oxygen species (ROS) production in the PMNs following phagocytosis of beads. In addition, levels of L-arginine and BH₄ were maintained or increased simultaneously to support the enzymatic activity of NOS in the bead activated PMNs. Moreover, translocation of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX) subunits along with iNOS was reconfirmed in the isolated phagosomes. We demonstrate that increase in the level of NO was supported by L-arginine and BH₄ to kill E. coli, by using PMNs from NOS2^?/? mice, human PMNs treated with biopterin inhibitor, N-acetyl serotonin (NAS), or by suspending human PMNs in L-arginine deficient medium. Altogether, this study demonstrates that following phagocytosis, sustained. NO production in the PMNs was well-maintained by redox sensitive cofactor, BH₄ and substrate, and L-arginine to enable microbial killing. Further results suggest NO production in the human PMNs, along with ROS and myeloperoxidase (MPO) is important to execute antimicrobial activity.     

Extracellular Secretion of Phytase from Transgenic Wheat Roots Allows Utilization of Phytate for Enhanced Phosphorus Uptake

A significant portion of organic phosphorus comprises of phytates which are not available to wheat for uptake. Hence for enabling wheat to utilize organic phosphorus in form of phytate, transgenic wheat expressing phytase from Aspergillus japonicus under barley root-specific promoter was developed. Transgenic events were initially screened via selection media containing BASTA, followed by PCR and BASTA leaf paint assay after hardening. Out of 138 successfully regenerated T_o events, only 12 had complete constructs and thus further analyzed. Positive T1 transgenic plants, grown in sand, exhibited 0.08–1.77, 0.02–0.67 and 0.44–2.14 fold increase in phytase activity in root extracts, intact roots and external root solution, respectively, after 4 weeks of phosphorus stress. Based on these results, T2 generation of four best transgenic events was further analyzed which showed up to 1.32, 56.89, and 15.40 fold increase in phytase activity in root extracts, intact roots and external root solution, respectively, while in case of real-time PCR, maximum fold increase of 19.8 in gene expression was observed. Transgenic lines showed 0.01–1.18 fold increase in phosphorus efficiency along with higher phosphorus content when supplied phytate or inorganic phosphorus than control plants. Thus, this transgenic wheat may aid in reducing fertilizer utilization and enhancing wheat yield.     

Leishmanicidal Triterpenes from Lantana camara

Two new natural triterpenes, lantaninilic acid and lantoic acid, along with the known triterpenes lantadene A, and oleanolic, ursolic, betulinic, lantanolic, and camaric acid, were obtained from the aerial parts of Lantana camara through bioassay‐guided isolation, monitoring the in vitro antileishmanial activity against promastigotes of Leishmania major. Oleanolic acid ( 3 ), ursolic acid ( 4 ), lantadene A ( 5 ), and lantanilic acid ( 7 ) showed significant leishmanicidal activities with IC₅₀ values of 53.0, 12.4, 20.4, and 21.3 μM , respectively. The IC₅₀ value of ursolic acid ( 4 ; 12.4 μM ) was found to be comparable with that of the standard drugs, pentamidine (IC₅₀ 15.0 μM ) and amphotericin B (IC₅₀ 0.31 μM ). The in vitro activities of L. camara and its constituents against promastigotes of Leishmania major are reported here for the first time.     

Investigating the interaction of anticancer drug temsirolimus with human transferrin: Molecular docking and spectroscopic approach

In our present study, binding between an important anti renal cancer drug temsirolimus and human transferrin (hTF) was investigated employing spectroscopic and molecular docking approach. In the presence of temsirolimus, hyper chromaticity is observed in hTF in UV spectroscopy suggestive of complex formation between hTF and temsirolimus. Fluorescence spectroscopy revealed the occurrence of quenching in hTF in the presence of temsirolimus implying complex formation taking place between hTF and temsirolimus. Further, the mode of interaction between hTF and temsirolimus was revealed to be static by fluorescence quenching analysis at 3 different temperatures. Binding constant values obtained employing fluorescence spectroscopy depicts strong interaction between hTF and temsirolimus; temsirolimus binds to hTF at 298 K with a binding constant of .32 × 10⁴ M^?1 implying the strength of this interaction. The negative Gibbs free energy obtained through quenching experiments is evident of the fact that the binding is spontaneous. CD spectra of hTF also showed a downward shift in the presence of temsirolimus as compared with free hTF implying complex formation between hTF and temsirolimus. Molecular docking was performed with a view to find out which residues are key players in this interaction. The importance of our study stems from the fact it will provide an insight into binding pattern of commonly administered renal cancer drug with an important protein that plays a pivotal role in many physiological processes.     

Glycation promotes the formation of genotoxic aggregates in glucose oxidase

This study investigates the effect of pentose sugars (ribose and arabinose) on the structural and chemical modifications in glucose oxidase (GOD) as well as genotoxic potential of this modified form. An intermediate state of GOD was observed on day 12 of incubation having CD minima peaks at 222 and 208?nm, characteristic of α-helix and a few tertiary contacts with altered tryptophan environment and high ANS binding. All these features indicate the existence of molten globule state of the GOD with ribose and arabinose on day 12. GOD on day 15 of incubation forms β structures as revealed by CD and FTIR which may be due to its aggregation. Furthermore, GOD on day 15 showed a remarkable increase in Thioflavin T fluorescence at 485?nm. Comet assay of lymphocytes and plasmid nicking assay in presence of glycated GOD show DNA damage which confirmed the genotoxicity of advance glycated end products. Hence, our study suggests that glycated GOD results in the formation of aggregates and the advanced glycated end products, which are genotoxic in nature.     

Iso-lines and inbred-lines confirmed loci that underlie resistance from cultivar ‘Hartwig’ to three soybean cyst nematode populations

Soybean [Glycine max (L.) Merr.] cultivars varied in their resistance to different populations of the soybean cyst nematode (SCN), Heterodera glycines, called HG Types. The rhg1 locus on linkage group G was necessary for resistance to all HG types. However, the loci for resistance to H. glycines HG Type 1.3- (race 14) and HG Type 1.2.5- (race 2) of the soybean cyst nematode have varied in their reported locations. The aims were to compare the inheritance of resistance to three nematode HG Types in a population segregating for resistance to SCN and to identify the underlying quantitative trait loci (QTL). ‘Hartwig’, a soybean cultivar resistant to most SCN HG Types, was crossed with the susceptible cultivar ‘Flyer’. A total of 92 F5-derived recombinant inbred lines (RILs; or inbred lines) and 144 molecular markers were used for map development. The rhg1 associated QTL found in earlier studies were confirmed and shown to underlie resistance to all three HG Types in RILs (Satt309; HG Type 0, P = 0.0001 R ² = 22%; Satt275; HG Type 1.3, P = 0.001, R ² = 14%) and near isogeneic lines (NILs; or iso-lines; Satt309; HG Type 1.2.5-, P = 0.001 R ² = 24%). A new QTL underlying resistance to HG Type 1.2.5- was detected on LG D2 (Satt574; P = 0.001, R ² = 11%) among 14 RILs resistant to the other HG types. The locus was confirmed in a small NIL population consisting of 60 plants of ten genotypes (P = 0.04). This QTL (cqSCN-005) is located in an interval previously associated with resistance to both SDS leaf scorch from ‘Pyramid’ and ‘Ripley’ (cqSDS-001) and SCN HG Type 1.3- from Hartwig and Pyramid. The QTL detected will allow marker assisted selection for multigenic resistance to complex nematode populations in combination with sudden death syndrome resistance (SDS) and other agronomic traits.     

Development of Organically Complexed-Bioaugmented Boron-Coated DAP and Its Effect on Yield and Quality of Canola (Brassica napus L.)

Journal of Plant Growth Regulation - Many agricultural soils fail to supply sufficient boron (B) and phosphorus (P) to growing plants due to their adsorption, precipitation and fixation phenomena....     

Synthesis and antibacterial activity of substituted flavones, 4-thioflavones and 4-iminoflavones

Synthesis of flavones, 4-thioflavones and 4-iminoflavones was carried out with the substitution of variable halogens, methyl, methoxy and nitro groups in the A, B and AB rings of the respective compounds and we also report here their antibacterial activity. Most of the synthesized compounds were found to be active against Escherichia coli, Bacillus subtilis, Shigella flexnari, Salmonella aureus, Salmonella typhi and Pseudomonas aeruginosa. Activity of 4-thioflavones and 4-iminoflavones was found to be higher than that of their corresponding flavone analogues. Investigated compounds having substituents like F, OMe and NO2 at 4'-position in ring-B exhibited enhanced activity and the presence of electronegative groups in the studied compounds showed a direct relationship to the antibacterial activity.