Home About us Contact
|
Home About us Contact | ||
|
|
||
Adenosyl Methionine (adenosyl + methionine)
Selected AbstractsS -Adenosyl methionine/S -adenosyl- L -homocysteine ratio determination by capillary electrophoresis employed as a monitoring tool for the antiviral effectiveness of adenosine analogsELECTROPHORESIS, Issue 10-11 2004Elena Sbrana Abstract S -Adenosyl- L -homocysteine hydrolase (SAHh) inhibitors have long been used as broad-range antivirals and have been recently evaluated as an experimental therapy of filovirus infections. In response to the need for a rapid laboratory testing method that could assess antiviral potency in vivo, our group developed a capillary electrophoresis (CE) method for the determination of the S -adenosyl- L -homocysteine (SAH) to S -adenosyl- L -methionine (SAM) ratio. After chloroacetaldehyde derivatization, SAH and SAM were detected using laser-induced fluorescence detection with a HeCd laser. Separation and quantitation of both SAH and SAM in human plasma were achieved in less than 1 min. The proposed method is rapid and reliable, and could be easily applied to routine monitoring of clinical and preclinical trials subjects. [source] High-phosphate-induced calcification is related to SM22, promoter methylation in vascular smooth muscle cellsJOURNAL OF BONE AND MINERAL RESEARCH, Issue 9 2010Addy Montes de Oca Abstract Hyperphosphatemia is closely related to vascular calcification in patients with chronic kidney disease. Vascular smooth muscle cells (VSMCs) exposed to high phosphate concentrations in vitro undergo phenotypic transition to osteoblast-like cells. Mechanisms underlying this transdifferentiation are not clear. In this study we used two in vitro models, human aortic smooth muscle cells and rat aortic rings, to investigate the phenotypic transition of VSMCs induced by high phosphate. We found that high phosphate concentration (3.3,mmol/L) in the medium was associated with increased DNA methyltransferase activity and methylation of the promoter region of SM22,. This was accompanied by loss of the smooth muscle cell,specific protein SM22,, gain of the osteoblast transcription factor Cbfa1, and increased alkaline phosphatase activity with the subsequent in vitro calcification. The addition of a demethylating agent (procaine) to the high-phosphate medium reduced DNA methyltransferase activity and prevented methylation of the SM22, promoter, which was accompanied by an increase in SM22, expression and less calcification. Additionally, downregulation of SM22,, either by siRNA or by a methyl group donor (S -adenosyl methionine), resulted in overexpression of Cbfa1. In conclusion, we demonstrate that methylation of SM22, promoter is an important event in vascular smooth muscle cell calcification and that high phosphate induces this epigenetic modification. These findings uncover a new insight into mechanisms by which high phosphate concentration promotes vascular calcification. © 2010 American Society for Bone and Mineral Research [source] Quantitative Lipid Metabolomic Changes in Alcoholic Micropigs With Fatty Liver DiseaseALCOHOLISM, Issue 4 2009Angela M. Zivkovic Background:, Chronic ethanol consumption coupled with folate deficiency leads to rapid liver fat accumulation and progression to alcoholic steatohepatitis (ASH). However, the specific effects of alcohol on key liver lipid metabolic pathways involved in fat accumulation are unknown. It is unclear whether lipid synthesis, lipid export, or a combination of both is contributing to hepatic steatosis in ASH. Methods:, In this study we estimated the flux of fatty acids (FA) through the stearoyl-CoA desaturase (SCD), phosphatidylethanolamine- N -methyltransferase (PEMT), and FA elongation pathways in relation to liver triacylglycerol (TG) content in Yucatan micropigs fed a 40% ethanol folate-deficient diet with or without supplementation with S -adenosyl methionine (SAM) compared with controls. Flux through the SCD and PEMT pathways was used to assess the contribution of lipid synthesis and lipid export respectively on the accumulation of fat in the liver. Liver FA composition within TG, cholesterol ester (CE), phosphatidylethanolamine, and phosphatidylcholine classes was quantified by gas chromatography. Results:, Alcoholic pigs had increased liver TG content relative to controls, accompanied by increased flux through the SCD pathway as indicated by increases in the ratios of 16:1n7 to 16:0 and 18:1n9 to 18:0. Conversely, flux through the elongation and PEMT pathways was suppressed by alcohol, as indicated by multiple metabolite ratios. SAM supplementation attenuated the TG accumulation associated with alcohol. Conclusions:, These data provide an in vivo examination of liver lipid metabolic pathways confirming that both increased de novo lipogenesis (e.g., lipid synthesis) and altered phospholipid metabolism (e.g., lipid export) contribute to the excessive accumulation of lipids in liver affected by ASH. [source] Functional Characterization of the Recombinant N -Methyltransferase Domain from the Multienzyme Enniatin SynthetaseCHEMBIOCHEM, Issue 9 2007Till Hornbogen Dr. Abstract A 51 kDa fusion protein incorporating the N -methyltransferase domain of the multienzyme enniatin synthetase from Fusarium scirpi was expressed in Saccharomyces cerevisiae. The protein was purified and found to bind S -adenosyl methionine (AdoMet) as demonstrated by cross-linking experiments with 14C-methyl-AdoMet under UV irradiation. Cofactor binding at equilibrium conditions was followed by saturation transfer difference (STD) NMR spectroscopy, and the native conformation of the methyltransferase was assigned. STD NMR spectroscopy yielded significant signals for H2 and H8 of the adenine moiety, H1' of D -ribose, and SCH3 group of AdoMet. Methyl group transfer catalyzed by the enzyme was demonstrated by using aminoacyl- N -acetylcysteamine thioesters (aminoacyl-SNACs) of L -Val, L -Ile, and L -Leu, which mimic the natural substrate amino acids of enniatin synthetase presented by the enzyme bound 4,-phosphopantetheine arm. In these experiments the enzyme was incubated in the presence of the corresponding aminoacyl-SNAC and 14C-methyl-AdoMet for various lengths of time, for up to 30 min. N -[14C-Methyl]-aminoacyl-SNAC products were extracted with EtOAc and separated by TLC. Acid hydrolysis of the isolated labeled compounds yielded the corresponding N -[14C-methyl] amino acids. Further proof for the formation of N - 14C-methyl-aminoacyl-SNACs came from MALDI-TOF mass spectrometry which yielded 23,212 Da for N -methyl-valyl-SNAC, accompanied by the expected postsource decay (PSD) pattern. Interestingly, L -Phe, which is not a substrate amino acid of enniatin synthetase, also proved to be a methyl group acceptor. D -Val was not accepted as a substrate; this indicates selectivity for the L isomer. [source] | ||||||||||