Why is folate effective in preventing neural tube closure defects?

Neural tube defects (NTDs) originate from a failure of the embryonic neural tube to close. The pathogenesis of NTDs is largely unknown. Fortunately, adequate maternal folate application is known to reduce the risk of human NTDs. However, why folate reduces NTDs is largely unknown. The main cause for NTDs is the disturbance of the cell growth in the neuroepithelium. Of course, rapid cell growth needs enough synthesis of nuclei acids. Interestingly, folate is used as a source for the synthesis of nucleic acids. Furthermore, glycine cleavage system (GCS) is essential for the synthesis of nucleic acids from folate, and very strongly expressed in neuroepithelial cells, suggesting that these highly proliferating cells need enough synthesis of nuclei acids and high amounts of folate. Taken together, I speculate the following hypothesis; (1) The closure of the neural tube requires rapid growth of neuroepithelial cells. (2) High rates of nuclei acids synthesis are needed for the rapid growth. (3) GCS, which is requisite in nucleic acid synthesis from folate, is expressed very strongly and functions robustly in neuroepithelial cells. (4) Pregnant women require 5-10-fold higher amounts of folate compared to non-pregnant women. (5) So, folate-deficient situations are easy to occur in neuroepithelial cells, resulting in NTDs. (6) Thus, folate is effective to prevent NTDs.

New developments in probing and targeting protein acylation in malaria, leishmaniasis and African sleeping sickness.

Infections by protozoan parasites, such as Plasmodium falciparum or Leishmania donovani, have a significant health, social and economic impact and threaten billions of people living in tropical and sub-tropical regions of developing countries worldwide. The increasing range of parasite strains resistant to frontline therapeutics makes the identification of novel drug targets and the development of corresponding inhibitors vital. Post-translational modifications (PTMs) are important modulators of biology and inhibition of protein lipidation has emerged as a promising therapeutic strategy for treatment of parasitic diseases. In this review we summarize the latest insights into protein lipidation in protozoan parasites. We discuss how recent chemical proteomic approaches have delivered the first global overviews of protein lipidation in these organisms, contributing to our understanding of the role of this PTM in critical metabolic and cellular functions. Additionally, we highlight the development of new small molecule inhibitors to target parasite acyl transferases.

MraY-antibiotic complex reveals details of tunicamycin mode of action.

The rapid increase of antibiotic resistance has created an urgent need to develop novel antimicrobial agents. Here we describe the crystal structure of the promising bacterial target phospho-N-acetylmuramoyl-pentapeptide translocase (MraY) in complex with the nucleoside antibiotic tunicamycin. The structure not only reveals the mode of action of several related natural-product antibiotics but also gives an indication on the binding mode of the MraY UDP-MurNAc-pentapeptide and undecaprenyl-phosphate substrates.

In vivo biocompatibility evaluation of some Bidens tripartita extracts in rats.

UNLABELLED: Adaptogens represent a class of herbs frequently used as a unique and natural alternative medicine and herbal remedy for treating the many forms of stress and different other pathological conditions. Bidens tripartite, a flowering plant from the genus Bidens, family Compositae, subfamily Asteroideae was widely used in traditional medicine for its antiseptic, anti-inflammatory, antioxidant, astringent, diuretic, febrifuge, narcotic and sedative effects. Phytochemical analysis of this plant has revealed the presence of flavonoids, xanthophylls, volatile oil, acetylene and polyacetylene, sterols, aurones, chalcones, caffeine and tannins. AIM: The in vivo biocompatibility evaluation of two extracts from Bidens tripartita plant in rats. MATERIAL AND METHODS: The vegetable product used for the study was obtained after maceration and extraction in alcohol. Flower powder was dissolved in absolute chloroform, re-extracted and filtered. After a complete dryness the product was extracted by the addition of ethanol then evaporated. The chemical composition of the extracts was determined. The administered dose of Bidens tripartita retained was 1/20 of lethal dose 50 (LD50). The experiment was carried out on white male Wistar rats (200-250g) divided into 3 groups of 7 animals each treated intraperitoneally as follows: Group I (Control): distilled water 0.1ml/10g weight; Group II (coded BT-alcoholic): 200mg/kbw alcoholic Bidens tripartita extract; Group III (coded BT-aqueous): 250mg/kbw aqueous Bidens tripartita extract. The biocompatibility properties of alcoholic and aqueous extracts from Bidens tritartita were studied by assessing their effects on blood count and serum biochemical tests. The following immune parameters: phagocytic capacity of peripheral neutrophils (NBT test) and serum complement activity were also evaluated. The data were presented as +/- SD and significance was tested by SPSS for Windows version 13.0 and ANOVA method. Experimental protocol was implemented according to the recommendations of the University Committee for Research and Ethical Issues and guidelines of IASP Committee for Research and Ethical Issue. RESULTS: Laboratory analysis did not show significant differences on leucocyte formula (GOT, GPT and LDH) or immune parameters (phagocytic capacity of peripheral neutrophils and serum complement activity) between alcoholic and aqueous B. tripartita extracts and distilled water, elements suggesting a good in vivo biocompatibility. CONCLUSIONS: In our experimental conditions, the alcoholic extract and aqueous extract from B. tripartita determined similar immune responses as distilled water following intraperitoneal administration in rats, indicative of good in vivo biocompatibility.

Alteraciones clínicas y bioquímicas en ratas tratadas con dosis altas de vitamina A / Clinical and biochemical alterations in rats treated with high doses of vitamin A

En el presente trabajo se estudió el efecto de la administración intramuscular de 30.000, 50.000 y 100.000 UI de palmitato de vitamina A/día, durante 7 días, respectivamente, sobre la actividad enzimática hepática en 45 ratas Wistar machos, de 12 semanas de edad, con pesos entre 180 y 200 gramos. El grupo control estuvo integrado por 15 ratas Wistar sanas, con género, edad y peso similares a los animales tratados. El consumo de alimentos y de agua, y el peso de las ratas se determinó al finalizar el período experimental. Las ratas se examinaron en busca de manifestaciones clínicas de toxicidad. Al final el estudio, las ratas se sacrificaron bajo anestesia con éter y se tomaron muestras de tejido hepático para la determinación de la actividad enzimática. La administración de vitamina A en exceso incrementó de manera significativa (p menor que 0,05) el contenido hepático del retinol, determinó diversos y variados signos clínicos (tales como: anorexia, pérdida de peso, alopecia, conjuntivitis, hemorragias internas y externas, alteraciones cutáneas y muerte de los animales) e incrementó (p menor que 0,05) la actividad de las siguientes enzimas: alanina aminotransferasa, aspartato aminotransferasa, maltasa ácida (alfa-1,4-glucosidasa ácida), proteasas ácidas, lactato dehidrogenasa y fosfatasa alcalina mientras que las actividades de la glucosa-6-fosfatasa, glucógeno fosforilasa, alfa-amilasa, colinesterasa y arginasa disminuyeron (p menor que 0,05) al comparar con los controles no tratados. Estos cambios son proporcionales a las dosis inyectadas de vitamina A. En conclusión, nuestros resultados proporcionan evidencias que la administración de dosis altas de vitamina A a corto plazo determina diversos y variados signos clínicos y produce una marcada alteración de la actividad enzimática hepática.

In the present work the effect of intramuscular administration of 30.000, 50.000 and 100.000 IU of vitamin A palmitate daily for seven days, respectively, on the liver enzyme activity in 45 white male Wistar rats, aged 12 weeks and weighing 180-200 g, have been studied. The group control was integrated by 15 healthy rats with similar characteristics (strain, gender, age and weight) to treated animals. Food and water consumption and body weights were recorded at the end of the experimental period. Rats were observed for clinical signs of toxicity. At the end of the study, rats were sacrificed under ether anesthesia. Liver samples were taken for the determination of enzyme activity. Administration of excess of vitamin A produced a significant (p menor 0.05) increase in the content of liver vitamin A, determined diverse and variable clinical signs (such as, anorexia, loss of body weight, alopecia, conjunctivitis, external and internal hemorrhages, skin abnormalities and death) and increased (p menor que 0.05) the activity of the following enzymes: alanine aminotransferase, aspartate aminotransferase, acid maltase (acid alfa-1,4-glucosidase), acid proteases, lactate dehydrogenase and alkaline phosphatase while glucose-6-phosphatase, glycogen phosphorylase, alfa-amylase, cholinesterase and arginase decreased (p menor que 0.05) as compared with untreated controls. These changes depend on the doses given of vitamin A. In conclusion, our results provide evidence that short-term administration of high doses of vitamin A determined diverse and variable clinical signs and produces a marked alteration of activity of liver enzymes.

Alkylation of opioid receptors by 5'-naltrindole-isothiocyanate injected into the nucleus accumbens of rats: receptor selectivity and anatomical diffusion.

Subtypes of the delta opioid receptor (Oprd1) have been suggested based on pharmacology studies. However, these subtypes have not been confirmed biochemically using either receptor binding assays or molecular cloning. Naltrindole-5'-isothiocyanate (5'-NTII) is an irreversible opioid antagonist that appears to selectively inhibit the actions of a subset of delta opioid agonists in vivo, referred to as putative delta-2 agonists. The biochemical and anatomical selectivity of wash-resistant inhibition of binding of [(3)H]DAMGO (Oprm1), [(3)H]DPDPE (Oprd1, putative subtype 1 agonist), or [(3)H]deltorphin II (Oprd1, putative subytpe 2 agonist) in coronal sections was assessed using quantitative in vitro autoradiography following injection of 5'-NTII into the nucleus accumbens in rats. 5'-NTII decreased [(3)H]deltorphin II to a greater extent than the binding of the other two radioligands following administration of 0.05-2.5 nmol. The effects of 5'-NTII were largely confined to the nucleus accumbens; however, some loss in the ventral caudate was also noted. In contrast, administration of the nonselective opioid receptor alkylating antagonist beta-chlornaltexamine (beta-CNA) over a similar range of doses was found to be nonselective for either delta radioligand, and produced greater inhibition of Oprm1 relative to Oprd1 binding, consistent with the nonselective pharmacological activity of this antagonist. Although 5'-NTII inhibited [(3)H]deltorphin II binding to a greater extent, the binding of the other two radioligands was decreased over a similar range of doses. Absolute conclusions regarding the involvement of delta-2 opioid receptors in pharmacological or physiological effects based on studies with 5'-NTII should therefore be tempered, and for site-directed studies it would be best to employ doses of 0.5 nmol or lower.

Effect of antiepileptic drugs on plasma lipids, lipoprotein (a), and liver enzymes.

We conducted a study to assess the effect of phenobarbital, carbamazepine, and valproate on serum lipid profiles and lipoprotein (a) in 64 children with epilepsy (aged between 1 and 15 years) admitted to the child neurology outpatient clinic between July 2000 and July 2002. The children were separated as group 1 (18 children), treated with phenobarbital, 5 mg/kg/day; group 2 (22 children), treated with carbamazepine, 10 to 15 mg/kg/day; and group 3 (24 children), treated with sodium valproate, 20 mg/kg/day. Plasma lipoprotein (a), total cholesterol, triglycerides, low-density lipoprotein cholesterol, high-density lipoprotein cholesterol, apolipoprotein A and apolipoprotein B levels, and liver enzymes alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, and gamma-glutamyltransferase were determined before the initiation of the treatment and at 3, 6, and 12 months of the treatment period. The mean age of children in group 1 was significantly low compared with those in groups 2 and 3 (P <.05). The mean pretreatment lipid levels among the groups were not significantly increased. The mean lipoprotein (a) levels were significantly increased in all groups at 3, 6, and 12 months of the treatment period (P <.05). The increase in alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, total cholesterol, low-density lipoprotein cholesterol, and high-density lipoprotein cholesterol at 3, 6, and 12 months was statistically significant in group 1 (P <.05). The higher levels in lipoprotein (a) (mean > 30 mg/dL) were observed only in carbamazepine-treated patients at 6 and 12 months. The percentage of children with lipoprotein (a) levels over 30 mg/dL was 44%, 63%, and 33% in the phenobarbital-, carbamazepine-, and valproate-treated children, respectively. Antiepileptic drugs significantly increase the level of lipoprotein (a), which is a major risk factor for atherosclerosis, and also have variable effects on other lipid parameters. Lipoprotein (a) levels should be closely followed in patients receiving antiepileptic drugs. (J Child Neurol 2006;21:70-74).

Manipulation of prenyl chain length determination mechanism of cis-prenyltransferases.

The carbon backbones of Z,E-mixed isoprenoids are synthesized by sequential cis-condensation of isopentenyl diphosphate (IPP) and an allylic diphosphate through actions of a series of enzymes called cis-prenyltransferases. Recent molecular analyses of Micrococcus luteus B-P 26 undecaprenyl diphosphate (UPP, C55) synthase [Fujihashi M, Zhang Y-W, Higuchi Y, Li X-Y, Koyama T & Miki K (2001) Proc Natl Acad Sci USA98, 4337-4342.] showed that not only the primary structure but also the crystal structure of cis-prenyltransferases were totally different from those of trans-prenyltransferases. Although many studies on structure-function relationships of cis-prenyltransferases have been reported, regulation mechanisms for the ultimate prenyl chain length have not yet been elucidated. We report here that the ultimate chain length of prenyl products can be controlled through structural manipulation of UPP synthase of M. luteus B-P 26, based on comparisons between structures of various cis-prenyltransferases. Replacements of Ala72, Phe73, and Trp78, which are located in the proximity of the substrate binding site, with Leu--as in Z,E-farnesyl diphosphate (C15) synthase--resulted in shorter ultimate products with C(20-35). Additional mutation of F223H resulted in even shorter products. On the other hand, insertion of charged residues originating from long-chain cis-prenyltransferases into helix-3, which participates in constitution of the large hydrophobic cleft, resulted in lengthening of the ultimate product chain length, leading to C(60-75). These results helped us understand reaction mechanisms of cis-prenyltransferase including regulation of the ultimate prenyl chain-length.

Idraparinux and liver enzymes: observations from the PERSIST trial.

A potential influence of idraparinux--a synthetic analogue of the pentasaccharide sequence in heparins--on plasma liver enzyme levels was analysed in 37 patients suffering from deep vein thrombosis and participating in the PERSIST trial. Plasma gamma-glutamyl-transferase, aspartate aminotransferase and alanine aminotransferase were determined prior to enoxaparin treatment (screening), prior to randomization (baseline) and once weekly during the 12-week treatment period. Patients were initially treated with weight-adjusted enoxaparin for 4-7 days and then randomized to either idraparinux (2.5, 5, 7.5 or 10 mg) or warfarin. Gamma-glutamyl-transferase was significantly increased after administration of enoxaparin at the baseline visit (P = 0.004) and in week 2 (P = 0.009) to return to screening levels in week 3 for the remaining study period (all P > 0.05). Aspartate aminotransferase (P = 0.001) and alanine aminotransferase (P < 0.001) were significantly increased at the baseline visit and returned to screening values at week 2 for the remaining study period (all P > 0.05). There was no significant difference between the mean values of plasma liver enzymes of the four idraparinux groups and the warfarin group in all 13 measurements. We concluded that idraparinux in contrast to enoxaparin does not increase plasma liver enzymes significantly.

The neuroprotective antioxidant alpha-lipoic acid induces detoxication enzymes in cultured astroglial cells.

alpha-Lipoic acid (LA), an antioxidant with broad neuroprotective capacity, is thought to act by scavenging reactive oxygen species and stimulation of glutathione synthesis. LA shows structural resemblance to dithiolethiones, like anethole dithiolethione (ADT). ADT protects against oxidative damage, primarily by induction of phase II detoxication enzymes, in particular NAD(P)H:quinone oxidoreductase (NQO1) and glutathione-S-transferase (GST). Therefore, we investigated whether LA, like ADT, is capable also of inducing these protective enzymes. Our data show that LA, like ADT, induces a highly significant, time- and concentration dependent, increase in the activity of NQO1 and GST in C6 astroglial cells. The LA or ADT mediated induction of NQO1 was further confirmed by quantitative PCR and western blot analysis. This work for the first time unequivocally demonstrates LA mediated upregulation of phase II detoxication enzymes, which may highly contribute to the compounds' neuroprotective potential. Moreover, the data support the notion of a common mechanism of action of LA and ADT.

Effect of organosulfur compounds from garlic and cruciferous vegetables on drug metabolism enzymes.

The frequent consumption of cruciferous vegetables and garlic is associated with several health benefits. These foods contain organosulfur compounds that are known to affect the biotransformation of xenobiotics, and therefore can influence the toxicity and carcinogenicity of environmental chemicals. In this article, we review the effects of isothiocyanates and diallyl sulfide on xenobiotic metabolism and the enzymes involved in the process. Isothiocyanates and diallyl sulfide can modulate the levels of phase I and phase II drug-metabolizing enzymes by affecting the transcriptional rates of their genes, the turnover rates of specific mRNAs or enzymes, or the enzyme activity. These compounds are not general enzyme inhibitors or inducers. They elicit selectivity in their mode of action. Elucidating the mechanisms involved in the alteration of drug-metabolizing enzymes by isothiocyanates and diallyl sulfide will increase our understanding of their possible effects on the biotransformation of drugs as well as the potential beneficial or detrimental effects of these organosulfur compounds.

Chemoprotection by organosulfur inducers of phase 2 enzymes: dithiolethiones and dithiins.

One of the major mechanisms of protection against carcinogenesis, mutagenesis, and other forms of toxicity mediated by carcinogens is the induction of enzymes involved in their metabolism, particularly phase 2 enzymes such as glutathione S-transferases, UDP-glucuronosyl transferases, and quinone reductases. Animal studies indicate that induction of phase 2 enzymes is a sufficient condition for obtaining chemoprevention and can be achieved by administering any of a diverse array of naturally-occurring and synthetic chemopreventive agents. Alliaceous and cruciferous plants are rich in organosulfur compounds with inducer activity. Indeed, monitoring of enzyme induction has led to the recognition or isolation of novel, potent chemopreventive agents such as 1,2-dithiole-3-thiones, dithiins and the isothiocyanate sulforaphane. For example, oltipraz, a substituted 1,2-dithiole-3-thione originally developed as an antischistosomal agent, possesses chemopreventive activity against different classes of carcinogens targeting multiple organs. Mechanistic studies in rodent models for chemoprevention of aflatoxin B1 (AFB1)-induced hepatocarcinogenesis by oltipraz indicates that increased expression of phase 2 genes is of central importance, although inhibition of phase 1 activation of aflatoxin B1 can also contribute to protection. Exposure of rodents to 1,2-dithiole-3-thiones triggers nuclear accumulation of the transcription factor Nrf2 and its enhanced binding to the Antioxidant Response Element, leading to transcriptional activation of a score of genes involved in carcinogen detoxification and attenuation of oxidative stress. Nrf2-deficient mice fail to induce many of these genes in response to oltipraz and the impact of this genotype on the chemopreventive efficacy of dithiolethiones is currently under investigation. To test the hypothesis that enzyme induction is a useful strategy for chemoprevention in humans, three key elements are necessary: a candidate agent, an at-risk population and modulatable intermediate endpoints. Towards this end, a placebo-controlled, double blind clinical trial of oltipraz was conducted in residents of Qidong, P.R. China who are exposed to dietary aflatoxins and who are at high risk for the development of liver cancer. Oltipraz significantly enhanced excretion of a phase 2 product, aflatoxin-mercapturic acid, a derivative of the aflatoxin-glutathione conjugate, in the urine of study participants administered 125 mg oltipraz by mouth daily. Administration of 500 mg oltipraz once a week led to a significant reduction in the excretion of the primary oxidative metabolite of AFB1, aflatoxin M1, when measured shortly after drug administration. While this study highlighted the general feasibility of inducing phase 2 enzymes in humans, a longer term intervention is addressing whether protective alterations in aflatoxin metabolism can be sustained for extended periods of time in this high-risk population. Food-based approaches to chemoprotection, targeted both to the general population and high-risk individuals, offer many practical advantages compared to the use of pharmaceutical agents. Thus, identification and utilization of naturally-occurring organosulfur chemoprotectors including dithiins should be a high priority.

Carbohydrate deficient transferrin in alcoholic liver disease: mechanisms and clinical implications.

Carbohydrate-deficient transferrin (CDT) is now considered to be the most sensitive and specific biological marker of alcohol abuse. The mechanism by which chronic alcohol consumption causes an elevation of CDT levels in serum is discussed. The sensitivity and specificity of various test procedures are compared, with special emphasis on the impact of liver disease. Clinical applications are reviewed, including the utility of CDT as a marker of relapse in alcoholic patients, and the use of CDT for the systematic screening of drinking in vulnerable populations as part of a public health approach to alcoholism.

Aspectos moleculares de la resistencia a insecticidas / Molecular basic of insecticide resistance

El surgimiento de resistencia en poblaciones de insectos es uno de los efectos indeseables asociados al uso de insecticidas, y es un buen ejemplo del modo en que ocurren los procesos microevolutivos. En 1908 se documentó por primera vez la existencia de insectos resistentes a insecticidas. Ahora se conocen casos de resistencia en más de 500 especies de artrópodos. Los principales mecanismos que confieren resistencia a insecticidas son penetración cuticular reducida, metabolismo degradativo aumentado y reducción en la susceptibilidad de los sitios de acción. Los métodos de la biología molecular permiten identificar las bases moleculares de esos mecanismos. El propósito de este artículo es reseñar el conocimiento disponible acerca de la biología molecular de la resistencia a insecticidas: mutaciones puntuales en genes de acetilcolinesterasa (Drosophila melanogaster) y del receptor de GABA (varias especies), inserciones en genes de transferasas (D. melanogaster) y del citocromo P450 (D. melanogaster), amplificación de genes de esterasas (Myzus persicae y Culex pipiens / quinquefasciatus complex), cambios que afectan la expresión del gen del citocromo P450 (Musca domestica), y una mutación ligada al gen del canal de sodio dependiente de voltaje (M. domestica)

Aspectos moleculares de la resistencia a insecticidas / Molecular basic of insecticide resistance

El surgimiento de resistencia en poblaciones de insectos es uno de los efectos indeseables asociados al uso de insecticidas, y es un buen ejemplo del modo en que ocurren los procesos microevolutivos. En 1908 se documentó por primera vez la existencia de insectos resistentes a insecticidas. Ahora se conocen casos de resistencia en más de 500 especies de artrópodos. Los principales mecanismos que confieren resistencia a insecticidas son penetración cuticular reducida, metabolismo degradativo aumentado y reducción en la susceptibilidad de los sitios de acción. Los métodos de la biología molecular permiten identificar las bases moleculares de esos mecanismos. El propósito de este artículo es reseñar el conocimiento disponible acerca de la biología molecular de la resistencia a insecticidas: mutaciones puntuales en genes de acetilcolinesterasa (Drosophila melanogaster) y del receptor de GABA (varias especies), inserciones en genes de transferasas (D. melanogaster) y del citocromo P450 (D. melanogaster), amplificación de genes de esterasas (Myzus persicae y Culex pipiens / quinquefasciatus complex), cambios que afectan la expresión del gen del citocromo P450 (Musca domestica), y una mutación ligada al gen del canal de sodio dependiente de voltaje (M. domestica) (AU)

Regulation of heme polymerizing activity and the antimalarial action of chloroquine.

Mice infected with Plasmodium berghei served as donors of erythrocytes with a high level of parasitemia for the study of ferriprotoporphyrin IX (FP) polymerization. Six hours after treatment of these mice with 3 micromol of chloroquine per 25 g of body weight, there were significant losses of heme polymerase I (HPA I). For chloroquine-susceptible (CS) P. berghei, the rate of FP polymerization decreased from 541 +/- 42 (mean +/- standard deviation; n = 12) to 51 +/- 19 (n = 8) nmol of FP polymerized per h per ml of packed erythrocytes (normalized to represent 1,000 parasites per 1,000 erythrocytes). For chloroquine-resistant (CR) P. berghei, the rate decreased from 284 +/- 19 (n = 16) to 124 +/- 11 (n = 6) nmol per h per ml. The chloroquine-induced loss of HPA I was accompanied by the accumulation of unpolymerized FP in CS P. berghei but not in CR P. berghei, which is consistent with the hypothesis that FP mediates the antimalarial action of chloroquine. Quinine treatment partially reversed the effects of chloroquine in CS P. berghei but not in CR P. berghei. Cycloheximide treatment antagonized the effects of chloroquine in both lines of parasites. To explain these findings, we propose that chloroquine, quinine, and cycloheximide perturb a regulatory process for HPA I. Furthermore, we propose that when chloroquine engages its target in the regulatory process, it initiates a chain of events which culminates in increased production, accessibility, or reactivity of a regulator (inactivator) of HPA I.

Formylmethanofuran: tetrahydromethanopterin formyltransferase from Methanopyrus kandleri - new insights into salt-dependence and thermostability.

BACKGROUND: Formylmethanofuran: tetrahydromethanopterin formyltransferase (Ftr) from the methanogenic Archaeon Methanopyrus kandleri (optimum growth temperature 98 degrees C) is a hyperthermophilic enzyme that is absolutely dependent on the presence of lyotropic salts for activity and thermostability. The enzyme is involved in the pathway of carbon dioxide reduction to methane and catalyzes the transfer of formyl from formylmethanofuran to tetrahydromethanopterin. RESULTS: The crystal structure of Ftr, determined to a resolution of 1:73 AE reveals a homotetramer composed essentially of two dimers. Each subunit is subdivided into two tightly associated lobes both consisting of a predominantly antiparallel beta sheet flanked by alpha helices forming an alpha/beta sandwich structure. The approximate location of the active site was detected in a region close to the dimer interface. CONCLUSIONS: The adaptation of Ftr against high lyotropic salt concentrations is structurally reflected by a large number of negatively charged residues and their high local concentration on the surface of the protein. The salt-dependent thermostability of Ftr might be explained on a molecular basis by ionic interactions at the protein surface, involving both protein and inorganic salt ions, and the mainly hydrophobic interactions between the subunits and within the core.

Zinc and liver cirrhosis: biochemical and histopathologic assessment.

Experimental liver cirrhosis was produced by administration of thioacetamide. Cirrhotic animals were divided into two groups: one group was given zinc sulphate and the second kept as cirrhotic control. Zinc-treated animals showed a restoration of normal hepatic and plasma zinc and copper levels. Similarly, plasma levels of aspartate aminotransferase, alanine aminotransferase, gamma-glutamyl aminotransferase, and total bilirubin decreased significantly. Light microscopic studies showed that most of the hepatocytes appeared normal in zinc-treated as compared with untreated cirrhotic animals. The amount of fibrin, reticulin, and collagen, which was high in the cirrhotic livers, decreased following zinc treatment. Staining with periodic acid Schiff's reagent showed the ability of hepatocytes to store glycogen after zinc treatment. These results revealed that zinc may have some beneficial effect in the treatment of liver cirrhosis.

Effect of long-term ethanol pretreatment on the metabolism of dichloromethane to carbon monoxide in rats.

The present study investigates the influence of long-term ethanol (ETOH) treatment of rats [10% (v/v) for 4, 12, and 36 weeks] on the metabolism of DCM after its oral and inhalative uptake to CO. Biotransformation of DCM to CO as measured by carboxyhemoglobin (COHb) formation was stimulated after long-term ETOH treatment in rats. A single oral dose of DCM (6.2 mmol/kg body mass) caused a significant increase of COHb, the maximum of about 9% occurring approximately 6 hr after DCM administration. In comparison to this control, in the blood of rats pretreated with ETOH (10% v/v) for 4, 12, and 36 weeks COHb values of 18, 17, and 13%, respectively, were measured. Long-term ETOH treatment followed by inhalation of 100, 500, and 2500 ppm DCM for 4 hr stimulated the formation of COHb, compared to controls. The elevation of COHb level was accompanied by decreased concentrations of DCM in the blood. The reason for the elevated biotransformation of DCM was ascertained by means of the determination of p-nitrophenol and aniline hydroxylation in liver microsomes of rats after long-term ETOH treatment to be an increase in cytochrome P450-dependent enzyme activities.

Farnesyltransferase as a target for anticancer drug design.

The currently understood function for Ras in signal transduction is in mediating the transmission of signals from external growth factors to the cell nucleus. Mutated forms of this GTP-binding protein are found in 30% of human cancers with particularly high prevalence in colon and pancreatic carcinomas. These mutations destroy the GTPase activity of Ras and cause the protein to be locked in its active, GTP bound form. As a result, the signaling pathways are activated, leading to uncontrolled tumor growth. Ras function in signaling requires its association with the plasma membrane. This is achieved by posttranslational farnesylation of a cysteine residue present as part of the CA1A2X carboxyl terminal tetrapeptide of all Ras proteins. The enzyme that recognizes and farnesylates the CA1A2X sequence, Ras farnesyltransferase (FTase), has become an important target for the design of inhibitors that might be interesting as antitumor agents. Several approaches have been taken in the search for in vivo active inhibitors of farnesyltransferase. These include the identification of natural products such as the chaetomellic and zaragozic acids that mimic farnesylpyrophosphate, bisubstrate transition state analogs combining elements of the farnesyl and tetrapeptide substrates and peptidomimetics that reproduce features of the carboxyl terminal tetrapeptide CA1A2X sequence. This last group of compounds has been most successful in showing highly potent inhibition of FTase and selective blocking of Ras processing in a range of Ras transformed tumor cell lines at concentrations as low as 10 nM. Certain peptidomimetics will also block tumor growth in various mouse models, with apparently few toxic side effects. These results suggest that farnesyltransferase inhibitors hold considerable promise as anticancer drugs in the clinic.