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Cellular and Molecular Life Sciences :... Jun 2009Cullin-RING E3 ubiquitin-Ligases (CRLs) are the most prominent class of ubiquitin-ligases. By controlling the stability of a cohort of key regulators, CRLs impinge on... (Review)
Review
Cullin-RING E3 ubiquitin-Ligases (CRLs) are the most prominent class of ubiquitin-ligases. By controlling the stability of a cohort of key regulators, CRLs impinge on many cellular and biological processes such as immunity, development, transcription, cell signalling and cell cycle progression. CRLs are multi-subunit complexes composed of a catalytic site and a substrate recognition module nucleated around a cullin scaffold protein. Most eukaryotic genomes encode at least five distinct cullins, and each of these cullins recruits a specific substrate-recognition module such that CRL complexes are modular. Despite their considerable diversity, CRLs are regulated by similar mechanisms. In particular, recent observations indicate that conformational variability induced by CRL dimerization and by conjugation of the ubiquitin-like protein NEDD8 on the cullin subunit stimulates substrate polyubiquitination. In this review, we discuss the composition of CRL complexes and the various molecular mechanisms controlling their activity.
Topics: Animals; Arabidopsis Proteins; Cullin Proteins; Drosophila Proteins; Enzyme Activation; NEDD8 Protein; Phosphorylation; Protein Binding; Protein Multimerization; Saccharomyces cerevisiae Proteins; Substrate Specificity; Ubiquitination; Ubiquitins
PubMed: 19194658
DOI: 10.1007/s00018-009-8712-7 -
Annales D'endocrinologie Dec 2002Energy exists as organic molecules and heat in living organisms. In adult mammals, body weight and fat content remain unchanged if energy intake is strictly equivalent... (Review)
Review
Energy exists as organic molecules and heat in living organisms. In adult mammals, body weight and fat content remain unchanged if energy intake is strictly equivalent to energy expenditure. In other words, regulation of body weight requires energy of foods to be entirely dissipated as heat. Imbalance between ingested energy and thermogenesis induces obesity or thinness. Alterations of food intake or energy expenditure represent the two causes of body weight disturbance. It is accepted that individuals differ in food efficiency i.e. ability to metabolize foods and store fat or totally burn nutrients. Mechanisms of food efficiency and futile cycles are presented. I started my research work analysing thermogenic mechanism in brown adipose tissue. Actually, in addition to white adipose tissue which is the major type of adipose tissue, mammals own another type of adipose tissue referred to as brown adipose tissue. This later tissue is an activatable thermogenic organ which oxidizes fatty acids and releases heat in blood stream. Brown fat is activated during exposure to the cold (in rodents), at birth, and during arousal in hibernators. My initial work helped to characterize a mitochondrial protein named uncoupling protein or UCP which is responsible for activation of fatty acid oxidation and heat production in brown adipocytes. Actually, in most cells, fifty per cent of oxidation energy is recovered as ATP in mitochondria through the process of coupling of respiration to ADP phosphorylation. In contrast to mitochondria of most tissues, brown adipocyte mitochondria can escape the obligatorily coupling of respiration and waste almost ninety per cent of respiration energy as thermogenesis. UCP characterization and its molecular cloning as well as antibodies obtention were used to better understand cellular thermogenesis. Brown adipocytes were identified in babies and adult patients with pheochromocytoma. More recently, research on the brown fat UCP helped us to identify UCP2, a UCP homolog present in most human and animal tissues. A family of UCPs exist in animals and plants. These UCPs may function as mitochondrial uncouplers. However, the ancient function of the UCPs may be rather associated to adaptation to oxygen and control of free radicals than to thermogenesis. Further studies of UCPs will improve the knowledge of mitochondrial metabolism and substrate oxidation. In other respects, analysis of molecular mechanisms controlling respiration uncoupling may contribute to new strategies of treatment of metabolic disorders such as obesity.
Topics: Adipose Tissue; Adipose Tissue, Brown; Animals; Body Composition; Body Weight; Carrier Proteins; Energy Intake; Energy Metabolism; Humans; Ion Channels; Membrane Proteins; Mitochondrial Proteins; Thermogenesis; Uncoupling Protein 1
PubMed: 12733325
DOI: No ID Found -
Current Drug Metabolism Feb 2008The placenta is a unique organ that is essential to a healthy and normal pregnancy. A number of phase I and II metabolizing enzymes are expressed at moderate levels in... (Review)
Review
The placenta is a unique organ that is essential to a healthy and normal pregnancy. A number of phase I and II metabolizing enzymes are expressed at moderate levels in the placenta, and have been proven to have the ability to metabolize certain xenobiotics. Depending on the substrate, this metabolic action may have significant clinical implications on how it affects the fetus. A wide variety of transporters including P-glycoprotein, breast cancer resistance protein, and multidrug resistance associated proteins have also been discovered in the placenta, and while most are found to have mainly physiological substrates, there are a number of xenobiotics which are also able to gain access to the fetus through transport across the placenta. Depending on the xenobiotics and its intended action, drug transport across the placenta may be desired, acceptable or undesirable. Medications administered to the mother but designed to work on the fetus are now being used increasingly, and demonstrates an important clinical implication in which drug transport across the placenta is desirable. However, medications designed to treat the mother but are also able to cross the placenta carry potential risks to damage the developing fetus, and it is therefore essential that the effects of different drugs on the fetus are known before they are administered during pregnancy. There is still much unknown about drug transport and drug metabolism in the placenta, and it is vital that in the future further research is done to discover the clinical implications of these activities in the placenta. This research is often complicated by the fact that it is unethical to run studies in pregnant women, and so research is often carried out in pregnant animals. These results are not always accurate, however, as the human's placental structure is different from the placenta in other animals. Drug metabolism and drug transport across the placenta should continue to be researched, and guidelines need to be developed to ensure that any medications used during pregnancy are safe to both the mother and the fetus, and that successful treatment of the medical condition is carried out.
Topics: ATP Binding Cassette Transporter, Subfamily B, Member 1; ATP Binding Cassette Transporter, Subfamily G, Member 2; ATP-Binding Cassette Transporters; Biological Transport; Humans; Membrane Transport Proteins; Multidrug Resistance-Associated Protein 2; Multidrug Resistance-Associated Proteins; Neoplasm Proteins; Organic Cation Transport Proteins; Pharmaceutical Preparations; Placenta; Serotonin Plasma Membrane Transport Proteins; Solute Carrier Family 22 Member 5
PubMed: 18288953
DOI: 10.2174/138920008783571828 -
Methods in Molecular Biology (Clifton,... 2015Despite the highly conserved helicase core, individual DEAD-box proteins are specialized in diverse RNA metabolic processes. One mechanism that determines DEAD-box...
Despite the highly conserved helicase core, individual DEAD-box proteins are specialized in diverse RNA metabolic processes. One mechanism that determines DEAD-box protein specificity is enzymatic regulation by other protein cofactors. In this chapter, we describe a protocol for purifying the Saccharomyces cerevisiae DEAD-box RNA helicase Dbp2 and RNA-binding protein Yra1 and subsequent analysis of helicase regulation. The experiments described here can be adapted to other RNA helicases and their purified cofactor(s).
Topics: DEAD-box RNA Helicases; Nuclear Proteins; Protein Binding; RNA-Binding Proteins; Saccharomyces cerevisiae Proteins
PubMed: 25579587
DOI: 10.1007/978-1-4939-2214-7_12 -
The Journal of Steroid Biochemistry and... May 2015CYP11A1 hydroxylates vitamin D3 producing 20S-hydroxyvitamin D3 [20(OH)D3] and 20S,23-dihydroxyvitamin D3 [20,23(OH)2D3] as the major and most characterized metabolites....
CYP11A1 hydroxylates vitamin D3 producing 20S-hydroxyvitamin D3 [20(OH)D3] and 20S,23-dihydroxyvitamin D3 [20,23(OH)2D3] as the major and most characterized metabolites. Both display immuno-regulatory and anti-cancer properties while being non-calcemic. A previous study indicated 20(OH)D3 can be metabolized by rat CYP24A1 to products including 20S,24-dihydroxyvitamin D3 [20,24(OH)2D3] and 20S,25-dihydroxyvitamin D3, with both producing greater inhibition of melanoma colony formation than 20(OH)D3. The aim of this study was to characterize the ability of rat and human CYP24A1 to metabolize 20(OH)D3 and 20,23(OH)2D3. Both isoforms metabolized 20(OH)D3 to the same dihydroxyvitamin D species with no secondary metabolites being observed. Hydroxylation at C24 produced both enantiomers of 20,24(OH)2D3. For rat CYP24A1 the preferred initial site of hydroxylation was at C24 whereas the human enzyme preferred C25. 20,23(OH)2D3 was initially metabolized to 20S,23,24-trihydroxyvitamin D3 and 20S,23,25-trihydroxyvitamin D3 by rat and human CYP24A1 as determined by NMR, with both isoforms showing a preference for initial hydroxylation at C25. CYP24A1 was able to further oxidize these metabolites in a series of reactions which included the cleavage of C23-C24 bond, as indicated by high resolution mass spectrometry of the products, analogous to the catabolism of 1,25(OH)2D3 via the C24-oxidation pathway. Similar catalytic efficiencies were observed for the metabolism of 20(OH)D3 and 20,23(OH)2D3 by human CYP24A1 and were lower than for the metabolism of 1,25(OH)2D3. We conclude that rat and human CYP24A1 metabolizes 20(OH)D3 producing only dihydroxyvitamin D3 species as products which retain biological activity, whereas 20,23(OH)2D3 undergoes multiple oxidations which include cleavage of the side chain.
Topics: Animals; Calcifediol; Dihydroxycholecalciferols; Humans; Hydroxylation; Oxidation-Reduction; Rats; Vitamin D; Vitamin D3 24-Hydroxylase
PubMed: 25727742
DOI: 10.1016/j.jsbmb.2015.02.010 -
Environmental Microbiology Sep 2015In metabolically versatile bacteria, carbon catabolite repression (CCR) facilitates the preferential assimilation of the most efficient carbon sources, improving growth...
In metabolically versatile bacteria, carbon catabolite repression (CCR) facilitates the preferential assimilation of the most efficient carbon sources, improving growth rates and fitness. In Pseudomonas putida, the Crc and Hfq proteins and the CrcZ and CrcY small RNAs, which are believed to antagonize Crc/Hfq, are key players in CCR. Unlike that seen in other bacterial species, succinate and glucose elicit weak CCR in this bacterium. In the present work, metabolic, transcriptomic and constraint-based metabolic flux analyses were combined to clarify whether P. putida prefers succinate or glucose, and to identify the role of the Crc protein in the metabolism of these compounds. When provided simultaneously, succinate was consumed faster than glucose, although both compounds were metabolized. CrcZ and CrcY levels were lower when both substrates were present than when only one was provided, suggesting a role for Crc in coordinating metabolism of these compounds. Flux distribution analysis suggested that, when both substrates are present, Crc works to organize a metabolism in which carbon compounds flow in opposite directions: from glucose to pyruvate, and from succinate to pyruvate. Thus, our results support that Crc not only favours the assimilation of preferred compounds, but balances carbon fluxes, optimizing metabolism and growth.
Topics: Bacterial Proteins; Carbon; Catabolite Repression; Gene Expression Regulation, Bacterial; Gluconeogenesis; Glucose; Glycolysis; Host Factor 1 Protein; Molecular Sequence Data; Pseudomonas putida; Pyruvic Acid; RNA, Small Untranslated; Repressor Proteins; Succinic Acid
PubMed: 25711694
DOI: 10.1111/1462-2920.12812 -
European Journal of Drug Metabolism and... 1982Absorption, tissue distribution, excretion, protein binding, and the metabolism of 14C labelled F-leurosine were studied in the rat. A triphasic curve for the...
Absorption, tissue distribution, excretion, protein binding, and the metabolism of 14C labelled F-leurosine were studied in the rat. A triphasic curve for the disappearance of the drug from blood was found. The bile was the major route of excretion: 80% of radioactivity was recovered in the bile during the first 28 h after i.v. administration. 54% of the F-leurosine binds, in the concentration range of 0.1 mumol/1-0.5 mmol/l to plasma proteins. The TLC and HPLC data suggested that the bile of rats administered [14C]-F-leurosine contained either none or only small amount of metabolic products, which were most probably due to chemical decomposition rather then metabolism. Rat liver homogenates did not metabolize [14C] F-leurosine to any detectable extent.
Topics: Animals; Blood Proteins; Intestinal Absorption; Kinetics; Male; Protein Binding; Rats; Rats, Inbred Strains; Tissue Distribution; Vinca Alkaloids
PubMed: 7067723
DOI: 10.1007/BF03189542 -
Journal of Clinical Pharmacy and... Jun 1998Liver disease is associated with reduced metabolic capacity for drugs that are metabolized by oxidative biotransformation. Three cytochrome P450 (P450 or CYP) gene... (Review)
Review
Liver disease is associated with reduced metabolic capacity for drugs that are metabolized by oxidative biotransformation. Three cytochrome P450 (P450 or CYP) gene families in liver microsomes (CYP 1, CYP2 and CYP3) appear to be responsible for much of the drug metabolism that takes place. The genetic polymorphism of the CYPs responsible for debrisoquine/ sparteine (CYP2D6) metabolism and S-mephenytoin (CYP2C19) metabolism has been well documented, but information on the impairment of each isoform in liver disease is still limited. There are two types of hepatic P450 function tests. One type consists of non-genetic P450 function tests (CYP1A2, 2A6, 2C9/10, 2E1 and 3A3/4), and probe drugs include caffeine, catalysed by CYP1A2, coumarin by CYP2A6, phenytoin by CYP2C6, chlorzoxazone by CYP2E1, and nifedipine, erythromycin and lidocaine by CYP3A3/4. The second type of genetic P450 function tests (CYP2C19 and CYP2D6) involves probe drugs such as S-mephenytoin, catalysed by CYP2C19, and debrisoquine and sparteine, catalysed by CYP2D6. The metabolism of the probe drugs used in non-genetic P450 function tests in patients with liver disease falls into two categories: reduced (CYP1A2, CYP2C, 2E1 and 3A) and unchanged (CYP2C). In genetic P450 function tests there seems to be a lesser degree of inhibition in poor metabolizers (PMs) than extensive metabolizers (EMs) among patients with liver disease. There have been very few reports on changes in metabolism of the probe drugs used in genetic P450 function tests in liver disease. In this paper the subject is reviewed.
Topics: Biotransformation; Cytochrome P-450 Enzyme System; Humans; Isoenzymes; Liver Diseases; Liver Function Tests
PubMed: 9831966
DOI: 10.1046/j.1365-2710.1998.00135.x -
The British Journal of Nutrition Aug 2012The nutritive value of food protein sources is dependent on the amino acid composition and the bioavailability of the nutritionally indispensable amino acids.... (Comparative Study)
Comparative Study Review
The nutritive value of food protein sources is dependent on the amino acid composition and the bioavailability of the nutritionally indispensable amino acids. Traditionally the methods developed to determine amino acid bioavailability have focused on intestinal absorption or digestibility, which is calculated as the percent of amino acid intake that does not appear in digesta or faeces. Traditional digestibility based methods do not always account for gut endogenous amino acid losses or absorbed amino acids which are unavailable due to the effect of heat processing and the presence of anti-nutritional factors, though methods have been developed to address these issues. Furthermore, digestibility based methods require the use of animal models, thus there is a need to develop in vivo methods that can be applied directly in human subjects to identify the proportion of dietary amino acids which is bioavailable, or metabolically available to the body for protein synthesis following digestion and absorption. The indicator amino acid oxidation (IAAO) method developed in our laboratory for humans has been systematically applied to determine almost all indispensable amino acid requirements in adult humans. Oxidation of the indicator amino acid is inversely proportional to whole body protein synthesis and responds rapidly to changes in the bioavailability of amino acids for metabolic processes. Using the IAAO concept, we developed a new in vivo method in growing pigs, pregnant sows and adult humans to identify the metabolic availability of amino acids in foods. The stable isotope based metabolic availability method is suitable for rapid and routine analysis in humans, and can be used to integrate amino acid requirement data with dietary amino acid availability of foods.
Topics: Amino Acids; Animals; Biological Availability; Diet; Dietary Proteins; Digestion; Female; Humans; Isotopes; Oxidation-Reduction; Pregnancy; Protein Biosynthesis
PubMed: 23107543
DOI: 10.1017/S0007114512002498 -
The Journal of Biological Chemistry Feb 2018Nicotinamide adenine dinucleotide (NAD) is an essential metabolite participating in cellular redox chemistry and signaling, and the complex regulation of NAD metabolism...
Nicotinamide adenine dinucleotide (NAD) is an essential metabolite participating in cellular redox chemistry and signaling, and the complex regulation of NAD metabolism is not yet fully understood. To investigate this, we established a NAD-intermediate specific reporter system to identify factors required for salvage of metabolically linked nicotinamide (NAM) and nicotinic acid (NA). Mutants lacking components of the NatB complex, and appeared as hits in this screen. NatB is an N-terminal acetyltransferase responsible for acetylation of the N terminus of specific Met-retained peptides. In NatB mutants, increased NA/NAM levels were concomitant with decreased NAD We identified the vacuolar pool of nicotinamide riboside (NR) as the source of this increased NA/NAM. This NR pool is increased by nitrogen starvation, suggesting NAD and related metabolites may be trafficked to the vacuole for recycling. Supporting this, increased NA/NAM release in NatB mutants was abolished by deleting the autophagy protein We next examined Tpm1 (tropomyosin), whose function is regulated by NatB-mediated acetylation, and Tpm1 overexpression () was shown to restore some NatB mutant defects. Interestingly, although largely suppressed NA/NAM release in NatB mutants, it did not restore NAD levels. We showed that decreased nicotinamide mononucleotide adenylyltransferase (Nma1/Nma2) levels probably caused the NAD defects, and was sufficient to restore NAD NatB-mediated N-terminal acetylation of Nma1 and Nma2 appears essential for maintaining NAD levels. In summary, our results support a connection between NatB-mediated protein acetylation and NAD homeostasis. Our findings may contribute to understanding the molecular basis and regulation of NAD metabolism.
Topics: Acetylation; Acetyltransferases; Autophagy-Related Proteins; Gene Deletion; Genes, Reporter; Homeostasis; Immunoprecipitation; Isoenzymes; Models, Molecular; Mutation; N-Terminal Acetyltransferase B; NAD; Nicotinamide-Nucleotide Adenylyltransferase; Protein Interaction Domains and Motifs; Protein Multimerization; Protein Processing, Post-Translational; Recombinant Proteins; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Tropomyosin
PubMed: 29317496
DOI: 10.1074/jbc.M117.807214