Moin Uddin

Methylglyoxal mediated conformational changes in histone H2A—generation of carboxyethylated advanced glycation end products

Methylglyoxal, an oxo-aldehyde has been implicated as a potential precursor in non enzymatic glycation reactions. Its role in the modification of extra cellular proteins has been extensively reported, but little is known about its modification of nuclear proteins, like histones. Here, we report the methylglyoxal induced modification of histone H2A which forms an essential part of intact core nucleosome. In this study commercially available histone H2A was subjected to in vitro non-enzymatic glycation by methylglyoxal. The structural alterations in the histone were characterised by various biophysical and biochemical techniques. The modified histone showed hyperchromicity at 276nm, loss in intrinsic tyrosine fluorescence intensity at 305nm along with a red shift, cross linking and dimer formation in SDS PAGE, induction of α-helix in CD spectroscopy, reduced hydrophobicity in ANS binding studies, accumulation of AGE products, increased carbonyl content, and appearance of a novel peak showing carboxyethylation complemented by a shift in amide I and amide II bands in ATR-FTIR spectroscopy. The modified histone exhibited increased melting temperatures (Tm) and enhanced heat capacities (Cp) in differential scanning calorimetric analysis. The results suggest that methylglyoxal significantly altered the structure of the nuclear histone H2A by non enzymatic glycation reaction. The conformational changes in histone H2A may influence the chromatin integrity which may have implications in various pathological conditions.

Role of peroxynitrite-modified H2A histone in the induction and progression of rheumatoid arthritis

Objective: Rheumatoid arthritis (RA) is an autoimmune disease of complex aetiology and pathogenesis. In recent years it has become evident that apoptotically modified histones exert a central role in the induction of autoimmunity, for example in systemic lupus erythematosus (SLE) and RA. Because nitric oxide (NO)-related species have been found in inflamed joints of arthritis patients, we investigated whether nitrotyrosine is present in sera of RA patients, and whether peroxynitrite-modified H2A histone is likely to be involved in the induction and progression of RA. Methods: Commercially available H2A histone was modified in vitro by peroxynitrite. The RA patients were divided into three groups on the basis of CRP, nitrite, total protein and IgG level. Sera of RA patients with high-titre rheumatoid factor (RF) were analysed for autoantibodies against native DNA and native and peroxynitrite-modified H2A histone. Binding characteristics and specificity of the autoantibodies were analysed by direct binding, inhibition enzyme-linked immunosorbent assay (ELISA), and band shift assay. Results: Sera from control subjects contained almost negligible amounts of 3-nitrotyrosine (3-NT); lower levels were found in group 1 RA patients in comparison to group 2 and group 3 patients, where the level of nitrotyrosine was progressively higher. Enzyme immunoassay data showed preferential binding of RA autoantibodies to peroxynitrite-modified H2A histone in comparison with native H2A histone or native DNA. The results suggest that peroxynitrite modification of self-antigen(s) can generate neoepitopes capable of inducing RA characteristic autoantibodies. Conclusion: The preferential binding of peroxynitrite-modified histones by autoantibodies derived from RA sera indicates a role for oxidatively modified and nitrated proteins in the initiation/progression of RA.

Autoimmune response to AGE modified human DNA: Implications in type 1 diabetes mellitus

Aims Non-enzymatic glycation of DNA both in vivo and in vitro results in generation of free radicals, known as glycoxidation. Glycoxidation leads to structural perturbation of DNA resulting in generation of neo-antigenic epitopes having implication in autoimmune disorders like diabetes mellitus. In this study human placental DNA was glycated with methylglyoxal (MG) and lysine (Lys) in the presence of Cu2+ and its auto-antibody binding was probed in Type 1 diabetes patients. Methods Glycation was carried out by incubating DNA with MG, Lys and Cu2+ for 24 h at 37 °C. Carboxyethyl deoxyguanosine (CEdG) formed in glycation reaction was studied by LC-MS and the pathway for Amadori formation was studied by ESI-MS techniques. Furthermore, binding characteristics of auto-antibodies in diabetes patients were assessed by direct binding, competitive ELISA and band shift assay. Results DNA glycation with MG, Lys and Cu2+ results in the formation of CEdG (marker of DNA glycation) which was confirmed by LC-MS. The intermediate stages of glycation were confirmed by ESI-MS technique. Serum from diabetes patients exhibited enhanced binding and specificity for glycated DNA as compared to native form. Conclusions Glycation of DNA has resulted in structural perturbation causing generation of neo-antigenic epitopes thus recognizing auto-antibodies in diabetes.

Use of N and P biofertilizers reduces inorganic phosphorus application and increases nutrient uptake, yield, and seed quality of chickpea

Leguminous crops suffer severely in soils poor in phosphorus. A 2-factor factorial experiment was conducted in a net-house to explore the effect of graded levels of P fertilizer (0, 30, and 60 kg P ha-1 or P0, P30, and P60, respectively) together with rhizobium (biological nitrogen fertilizer (BNF)) and/or phosphate-solubilizing bacteria (biological phosphorus fertilizer (BPF)) in terms of nutrient uptake, yield, and quality of chickpea (Cicer arietinum L.). Phosphorus was applied as basal dose, while seeds were treated with respective biofertilizer(s) before sowing according to the treatments [BF0 (control), BNF, BPF, and BNF+BPF]. Concerning the main effects, P60 proved superior or equivalent to P30, while among the biofertilizer treatments, BNF+BPF gave the greatest values for nutrient uptake as well as for yield and quality parameters. The interaction between P levels and biofertilizer treatments was generally significant. 30 kg P ha-1 applied with N and P biofertilizers (P30 × BNF+BPF) was the most profitable interaction for N uptake as well as for yield and quality characteristics. Compared to P60 applied alone (P60 × BF0), P30 × BNF+BPF resulted in greater N uptake (27.3%), seed yield (21.1%), and the content of seed protein (2.9%) and carbohydrate (5.6%). Furthermore, P30 × BNF+BPF was statistically equal to P60 × BNF+BPF with regard to N uptake, seed yield, and protein content as well as for most yield components. Thus, P30 × BNF+BPF saved 30 kg P ha-1 of the costly inorganic P fertilizer to achieve the greatest crop yield and quality.

Radiolytically depolymerized sodium alginate improves physiological activities, yield attributes and composition of essential oil of Eucalyptus citriodora Hook

Eucalyptus citriodora Hook. is highly valued for its citronellal-rich essential oil (EO) extracted from its leaves. Hence, escalated EO production of eucalyptus is the need of hour. Marine polysaccharides (sodium alginate) are processed through gamma radiation of particular intensity, to obtain the irradiated sodium alginate (ISA). A pot experiment was conducted to study the effect of foliar application of ISA on growth, biochemical, physiological, EO yield and composition of E. citriodora. The treatments were applied as: foliar spray of deionized water only (control), seed soaked with ISA (90 mg L(-1)) and foliar spray of ISA with 30, 60, 120 and 240 mg L(-1). The treatment 6 (spray of ISA at 120 mg L(-1)) showed the highest value for most of the parameters studied. It also enhanced the EO content (33.3%), EO yield (86.7%), citronellal content (63.4%) and citronellal yield (205.5%) as compared to the control.

Dicarbonyl Induced Structural Perturbations Make Histone H1 Highly Immunogenic and Generate an Auto-Immune Response in Cancer.

Increased oxidative stress under hyperglycemic conditions, through the interaction of AGEs with RAGE receptors and via activation of interleukin mediated transcription signalling, has been reported in cancer. Proteins modifications are being explored for their roles in the development and progression of cancer and autoantibody response against them is gaining interest as a probe for early detection of the disease. This study has analysed the changes in histone H1 upon modification by methylglyoxal (MG) and its implications in auto-immunopathogenesis of cancer. Modified histone showed modifications in the aromatic residues, changed tyrosine microenvironment, intermolecular cross linking and generation of AGEs. It showed masking of hydrophobic patches and a hypsochromic shift in the in ANS specific fluorescence. MG aggressively oxidized histone H1 leading to the accumulation of reactive carbonyls. Far UV CD measurements showed di-carbonyl induced enhancement of the alpha structure and the induction of beta sheet conformation; and thermal denaturation (Tm) studies confirmed the thermal stability of the modified histone. FTIR analysis showed amide I band shift, generation of a carboxyethyl group and N-Cα vibrations in the modified histone. LCMS analysis confirmed the formation of Nε-(carboxyethyl)lysine and electron microscopic studies revealed the amorphous aggregate formation. The modified histone showed altered cooperative binding with DNA. Modified H1 induced high titre antibodies in rabbits and the IgG isolated form sera of rabbits immunized with modified H1 exhibited specific binding with its immunogen in Western Blot analysis. IgG isolated from the sera of patients with lung cancer, prostate cancer, breast cancer and cancer of head and neck region showed better recognition for neo-epitopes on the modified histone, reflecting the presence of circulating autoantibodies in cancer. Since reports suggest a link between AGE-RAGE axis and carcinogenesis, glycoxidation of histone H1 and its immunogenicity paves ways for understanding role of glycoxidatively damaged nuclear proteins in cancer.

Effect of Co-60 gamma irradiated chitosan and phosphorus fertilizer on growth, yield and trigonelline content of Trigonella foenum-graecum L.

Abstract Natural marine polysaccharides such as chitosan, sodium alginate and carrageenan; after depolymerization by different techniques, have shown promising plant growth promoting and other biological activities. Chitosan after irradiation with Co-60 gamma-rays have proven to be a miraculous plant growth promoter in different plants. Trigonelline; an anti-diabetic compound is obtained mainly from the seeds of Fenugreek (Trigonella foenum-graecum L.). Trigonelline counteracts diabetes mellitus (DM) through the operation of the mechanisms of insulin secretion, modulation of β-cell regeneration and stimulation of activity of glucose metabolism related enzymes. Considering the medicinal importance a pot experiment was conducted to investigate the effect of foliar application of Co-60 gamma irradiated chitosan (IC) with soil applied phosphorus supplement on growth, biochemical and quality attributes of fenugreek. Four concentrations of irradiated chitosan were used (0, 40, 80 and 120mgL−1) individually as well as in combination with single dose of phosphorus 40kgha−1. Un-irradiated chitosan (UN) 40mgL−1 and de-ionized water were used as control. IC significantly affected almost all the parameters including seed yield, trigonelline content and trigonelline yield. There were 10 treatments and among all, P40+IC 40mgL−1 proved to be the best, which increased total alkaloid content by 34.9%, seed yield by 125.4%, and trigonelline content by 17.8%. Contents of photosynthetic pigments and activity of nitrate reductase, carbonic anhydrase enzymes were also significantly enhanced.

Modulation of physiological activities, active constituents and essential oil production of Mentha arvensis L. by concomitant application of depolymerised carrageenan, triacontanol and 28-homobrassinolide

Abstract Cornmint (Mentha arvensis L.) constitutes most important source of therapeutic agents used in the alternative systems of medicine. The essential oil of cornmint has wide applications in pharmaceutical, agrochemical and flavoring industries worldwide. This study was conducted to explore the individual as well as combined effect of the best foliar concentrations of gamma-irradiated carrageenan (IC), triacontanol (TRIA) and 28-homobrassinolide (HBR) on growth, yield and quality of cornmint. Foliar application of IC, TRIA and HBR, applied individually on plants, significantly improved the plant attributes studied. However, combined application of these plant growth regulators (PGRs) was more effective compared to their individual application. In comparison to other applied treatments and the control, the combination of three PGRs (80 ppm IC + 10−6 M TRIA + 10−7 M HBR) proved to be the best for most of the growth, physiological, biochemical and agronomic parameters studied. Combined application of the tested PGRs excelled the control in per plant yield of menthol, menthone and menthyl acetate by 135.9 and 134.1%, 180.0 and 161.1% and by 225.0 and 187.5% at 100 and 120 days after planting, respectively.

Radiolytically degraded sodium alginate enhances plant growth, physiological activities and alkaloids production in Catharanthus roseus L.

Abstract Catharanthus roseus (L.) G. Don (Family Apocynaceae) is a medicinal plant that produces indole alkaloids used in cancer chemotherapy. The anticancerous alkaloids, viz. vinblastine and vincristine, are mainly present in the leaves of C. roseus. High demand and low yield of these alkaloids in the plant has led to explore the alternative means for their production. Gamma irradiated sodium alginate (ISA) has proved as a plant growth promoting substance for various medicinal and agricultural crops. A pot culture experiment was carried out to explore the effect of ISA on plant growth, physiological activities and production of anticancer alkaloids (vinblastine and vincristine) in C. roseus at 120 and 150 days after planting (DAP). Foliar application of ISA (0, 20, 40, 60, 80 and 100mgL−1) significantly improved the performance of C. roseus. 80mgL−1 of ISA enhanced the leaf-yield by 25.3 and 30.2% and the herbage-yield by 29.4 and 34.4% at 120 and 150 DAP, respectively, as compared to the control. The spray treatment of ISA at 80mgL−1 improved the yield of vinblastine by 66.7 and 71.4% and of vincristine by 67.6 and 75.6% at 120 and 150 DAP, respectively, in comparison to the control. As compared to control, the application of ISA at 80mgL−1 resulted in the maximum swell in the content and yield of vindoline, increasing them by 18.9 and 20.8% and by 81.8 and 87.2% at 120 and 150 DAP, respectively.

Cumulative effect of gibberellic acid and phosphorus on crop productivity, biochemical activities and trigonelline production in Trigonella foenum-graecum L.

Abstract Fenugreek (Trigonella foenum-graecum L.) is an antidiabetic plant. Its bioactive compound, trigonelline, is known to counter diabetes through insulin secretion, modulation of β cell regeneration and quick activity of glucose metabolism related enzymes. A pot experiment was conducted in the natural conditions of net house of the Department of Botany, Aligarh Muslim University, Aligarh (UP), India, to evaluate the effect of four concentrations of GA3 (0, 10−7 M, 10−6 M and 10−5 M), alone and in combination with phosphorus (40 kg P ha−1), on growth, biochemical and yield attributes of fenugreek. Compared to control, the combination of GA3 and phosphorus (P40 + 10−6 M GA3) significantly increased the activities of nitrate reductase (30.8%) and carbonic anhydrase (30.7%) enzymes; it also enhanced the seed yield (140.6%) and the content of total chlorophyll (28.5%) and carotenoids (26%). There was also significant increase (19.51%) in the content of seed trigonelline.