Impact of short- and medium-chain organic acids, acylcarnitines, and acyl-CoAs on mitochondrial energy metabolism

Accumulation of organic acids as well as their CoA and carnitine esters in tissues and body fluids is a common finding in organic acidurias, beta-oxidation defects, Reye syndrome, and Jamaican vomiting sickness. Pathomechanistic approaches for these disorders have been often focused on the effect of accumulating organic acids on mitochondrial energy metabolism, whereas little is known about the pathophysiologic role of short- and medium-chain acyl-CoAs and acylcarnitines. Therefore, we investigated the impact of short- and medium-chain organic acids, acylcarnitines, and acyl-CoAs on central components of mitochondrial energy metabolism, namely alpha-ketoglutarate dehydrogenase complex, pyruvate dehydrogenase complex, and single enzyme complexes I-V of respiratory chain. Although at varying degree, all acyl-CoAs had an inhibitory effect on pyruvate dehydrogenase complex and alpha-ketoglutarate dehydrogenase complex activity. Effect sizes were critically dependent on chain length and number of functional groups. Unexpectedly, octanoyl-CoA was shown to inhibit complex III. The inhibition was noncompetitive regarding reduced ubiquinone and uncompetitive regarding cytochrome c. In addition, octanoyl-CoA caused a blue shift in the gamma band of the absorption spectrum of reduced complex III. This effect may play a role in the pathogenesis of medium-chain and multiple acyl-CoA dehydrogenase deficiency, Reye syndrome, and Jamaican vomiting sickness which are inherited and acquired conditions of intracellular accumulation of octanoyl-CoA.     

Role of curcumin in health and disease

Curcumin (diferuloylmethane) is an orange-yellow component of turmeric (Curcuma longa), a spice often found in curry powder. In recent years, considerable interest has been focused on curcumin due to its use to treat a wide variety of disorders without any side effects. It is one of the major curcuminoids of turmeric, which impart its characteristic yellow colour. It was used in ancient times on the Indian subcontinent to treat various illnesses such as rheumatism, body ache, skin diseases, intestinal worms, diarrhoea, intermittent fevers, hepatic disorders, biliousness, urinary discharges, dyspepsia, inflammations, constipation, leukoderma, amenorrhea, and colic. Curcumin has the potential to treat a wide variety of inflammatory diseases including cancer, diabetes, cardiovascular diseases, arthritis, Alzheimer's disease, psoriasis, etc, through modulation of numerous molecular targets. This article reviews the use of curcumin for the chemoprevention and treatment of various diseases.     

Heparinase selectively sheds heparan sulphate from the endothelial glycocalyx

A healthy vascular endothelium is coated by the endothelial glycocalyx. Its main constituents are transmembrane syndecans and bound heparan sulphates. This structure maintains the physiological endothelial permeability barrier and prevents leukocyte and platelet adhesion, thereby mitigating inflammation and tissue oedema. Heparinase, a bacterial analogue to heparanase, is known to attack the glycocalyx. However, the exact extent and specificity of degradation is unresolved. We show by electron microscopy, immunohistological staining and quantitative measurements of the constituent parts, that heparinase selectively sheds heparan sulphate from the glycocalyx, but not the syndecans.     

The inhibitory effects of acidification and augmented oxygen pressure on water oxidation

Cyanobacteria, algae, and plants produce dioxygen from water. Driven and clocked by light quanta, the catalytic Mn(4)Ca Tyrosine centre accumulates four oxidizing equivalents before it abstracts four electrons from water and liberates dioxygen and protons. Intermediates of this reaction cascade are short-lived (<100 micros) and difficult to detect. By application of high oxygen pressure to cyanobacterial PSII-core-complexes, we have previously suppressed the transition from the highest oxidation state of the centre to the lowest by stabilizing a (peroxy) intermediate. Here, we investigated the inhibitory interplay of acidification and augmented oxygen pressure. Starting from pH 6.5, acidification increasingly inhibited the reduction of the highest oxidized state and resulted in a lower oxygen partial pressure for half inhibition. Oxygen and proton interfere with different steps of the reaction cascade.     

Polymerized coumaric acid as a model substrate for terrestrial-derived dissolved organic carbon utilized by aquatic microorganisms

It is now widely accepted that many surface waters receive more terrestrial carbon than assumed in the past, and that aquatic food webs are largely based on the supply of external dissolved organic carbon. However, very little information is available on how efficiently external carbon is utilized by microorganisms and transported to consumers of higher trophic levels. To address this issue, we prepared and tested polymers of ¹⁴C-p-coumaric acid (PCA) as a model substrate for terrestrial organic carbon. Photodegradation products that can be considered potential substrates for microorganisms were identified using hyphenated techniques, including gas chromatography-mass spectrometry (GC/MS) and ion chromatography-electrospray ionization mass spectrometry (IC/MS). Photolysis of PCA released monomeric phenol derivatives, e.g. 4-hydroxybenzaldehyde. The photolysis products observed were similar to those characteristic for natural organic carbon. Both a heterotrophic bacteria assemblage and a cultured algae strain exhibiting heterotrophic capabilities proved capable of utilizing the model substrate. Irradiation of PCA increased the uptake rate approximately eight times for the bacteria, but no significant increase was observed for the algae. Potential sources of interferences, e.g. the uptake of ¹⁴CO₂ released by photolysis, were addressed. It was concluded that PCA is a suitable substrate to study the metabolism of terrestrial DOC within aquatic communities.     

Cloning,expression and characterisation of CYP102A7, a self-sufficient P450 monooxygenase from <Emphasis Type="Italic">Bacillus licheniformis</Emphasis>

Cytochrome P450 monooxygenases of the CYP102A subfamily are single-component natural fusion proteins consisting of a heme domain and a diflavin reductase. The characterised CYP102A enzymes are fatty acid hydroxylases with turnover rates of several thousands per minute. In search of new P450s with similar activities, but with a broader substrate spectrum, we cloned, expressed and characterised CYP102A7 from Bacillus licheniformis. As expected, CYP102A7 was active towards medium-chain fatty acids but showed a strong preference for saturated over unsaturated fatty acids, which could not be observed for either of the CYP102A members so far. Besides fatty acids, CYP102A7 was able to catalyse the oxidation of cyclic and acyclic terpenes with high activity and coupling efficiency. For example, (R)-(+)-limonene was converted with activity of 220 nmol nmol P450(-1) min(-1) and 80% coupling. Unusual for enzymes of the CYP102A subfamily was the deethylation activity of CYP102A7 towards 7-ethoxycoumarin. Furthermore, this monooxygenase, though having a moderate thermal stability, exhibited 50% of its initial activity in the presence of 26% DMSO. Comparison of the homology models of CYP102A7 and other members of the CYP102A subfamily revealed distinct differences in the shape of the substrate access channel and the active site, which might explain differences in catalytic properties of these homologous enzymes.     

Quinolone alkaloids from Waltheria douradinha

A phytochemical investigation of the stems of Waltheria douradinha resulted in isolation of two 4-quinolone alkaloids, waltherione B and vanessine, along with three known alkaloids, waltherione A, antidesmone and O-methyltembamide. Their structures were elucidated on the basis of their 2D NMR spectroscopic analyses, and from X-ray crystallographic analysis of waltherione A and the O-methyl derivative of waltherione B. Additionally, waltherione B and vanessine, and the O- and N-methyl derivatives of waltherione A and waltherione B, were evaluated for their antimicrobial activities; only vanessine displayed any (weak) antimicrobial activity.     

Teaching (and learning from) metabolomics: the 2006 PlantMetaNet ETNA Metabolomics Research School

Under the auspices of the European Training and Networking Activity programme of the European Union, a 'Metabolic Profiling and Data Analysis' Plant Genomics and Bioinformatics Summer School was hosted in Potsdam, Germany between 20 and 29 September 2006. Sixteen early career researchers were invited from the European Union partner nations and the so-called developing nations ( Appendix ). Lectures from invited leading European researchers provided an overview of the state of the art of these fields and seeded discussion regarding major challenges for their future advancement. Hands-on experience was provided by an example experiment – that of defining the metabolic response of Arabidopsis to treatment of a commercial herbicide of defined mode of action. This experiment was performed throughout the duration of the course in order to teach the concepts underlying extraction and machine handling as well as to provide a rich data set with which the required computation and statistical skills could be illustrated. Here we review the state of the field by describing both key lectures given at and practical aspects taught at the summer school. In addition, we disclose results that were obtained using the four distinct technical platforms at the different participating institutes. While the effects of the chosen herbicide are well documented, this study looks at a broader number of metabolites than in previous investigations. This allowed, on the one hand, not only to characterise further effects of the herbicide than previously observed but also to detect molecules other than the herbicide that were obviously present in the commercial formulation. These data and the workshop in general are all discussed in the context of the teaching of metabolomics.  

  Appendix.   List of participants in the school     

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