Showing papers on "Lanosterol published in 1985"
TL;DR: SF 86-327 is a new antimycotic agent of the allylamine type that appears to be the inhibition of ergosterol biosynthesis at the point of squalene epoxidation, as was previously found with the related compound naftifine.
Abstract: SF 86-327 is a new antimycotic agent of the allylamine type. Its primary action appears to be the inhibition of ergosterol biosynthesis at the point of squalene epoxidation, as was previously found with the related compound naftifine. Biosynthesis was measured by incorporation of [14C]acetate into sterols in cells of Candida albicans, Candida parapsilosis, Torulopsis glabrata, and the dermatophyte Trichophyton mentagrophytes. There was a positive correlation between the SF 86-327 concentrations needed for inhibition of growth and of sterol synthesis in these four fungi. The greater antifungal efficacy of SF 86-327 in comparison with naftifine was also reflected in the relative activities of the two compounds as sterol synthesis inhibitors. Inhibition was maximal at neutral pH. A similar degree of inhibition was found in cell-free extracts when [14C]mevalonate was used as substrate. In all cases, inhibition of sterol synthesis was accompanied by a parallel accumulation of labeled squalene. SF 86-327 and naftifine had no significant effect on initial enzymes of the ergosterol pathway, measured by incorporation of [14C]acetyl coenzyme A, or on steps distal to squalene epoxidation, measured by conversion of labeled squalene 2,3-epoxide or lanosterol. Both allylamines were highly selective for fungal, as opposed to mammalian, sterol biosynthesis. SF 86-327 caused slight inhibition of squalene epoxidation in a rat liver cell-free system, but at concentrations three to four orders of magnitude greater than those required for inhibition of the fungal pathway.
155 citations
TL;DR: The present investigation was conducted to compare homogeneous preparations of FABP and SCP-2 with respect to their capacities to participate as carrier proteins in reactions involving sterols or fatty acids, and shows that they have separate and distinct physiological functions.
110 citations
TL;DR: The chromatographic and spectrometric sterol analyses and the isotopic tracer findings suggested that ketoconazole impaired the cytochrome P-450 dependent 14 alpha-demethylation of lanosterol, that cholesterol was neither biosynthesized nor metabolized, and that the physiological functions of 5-dehydroepisterol had sterol structural requirements not entirely met by cholesterol.
71 citations
TL;DR: Results show that SCP2 enhances 7 alpha-hydroxycholesterol formation by rat liver microsomes (microsomal fractions), utilizing either endogenous membrane cholesterol, cholesterol supplied exogenously in serum or in the form of cholesterol/phospholipid liposomes.
Abstract: Sterol carrier protein2 (SCP2) is known to stimulate utilization of cholesterol in enzymic reactions in which cholesterol is the substrate. Substantial recent experimental evidence indicates that SCP2: activates enzymic conversion of intermediates between lanosterol and cholesterol; stimulates the microsomal conversion of cholesterol into cholesterol ester in rat liver; and enhances mitochondrial utilization of cholesterol for pregnenolone formation in the adrenals. The conversion of cholesterol into 7 alpha-hydroxycholesterol is the rate-limiting step in bile-acid synthesis. We therefore investigated the effect of SCP2 on this physiologically critical reaction by using a gas-chromatography-mass-spectrometry procedure that measures the mass of 7 alpha-hydroxycholesterol formed. The results show that SCP2 enhances 7 alpha-hydroxycholesterol formation by rat liver microsomes (microsomal fractions), utilizing either endogenous membrane cholesterol, cholesterol supplied exogenously in serum or in the form of cholesterol/phospholipid liposomes. Microsomes immunotitrated with anti-SCP2 antibody exhibited considerably less capacity to synthesize 7 alpha-hydroxycholesterol, which was restored to control levels on addition of purified SCP2. These data are consistent with the suggestion that SCP2 may be of physiological significance in the overall metabolism of cholesterol.
58 citations
TL;DR: The 14- methyl group prevents lanosterol from causing the increase in motional order of the phospholipid hydrocarbon chains characteristic of cholesterol, and suggests little disorganization of the membrane structure due to the 14-methyl group.
54 citations
TL;DR: Non-specific lipid transfer protein (nsL-TP) purified from rat and bovine liver accelerates the transfer of all common diacylglycerophospholipids, cholesterol as well as glycosphingolipid and gangliosides between membranes and is capable of mediating net transfer of cholesterol to membranes.
54 citations
TL;DR: The results show that at least some of the biochemical processes of Acanthamoeba polyphaga might be phylogenetically related to those of unicellular algae.
Abstract: The soil amoeba Acanthamoeba polyphaga is capable of synthesizing its sterols de novo from acetate. The major sterols are ergosterol and poriferasta-5,7,22-trienol. Furthermore C28 and C29 sterols of still unknown structure with an aromatic B-ring are also synthesized by the amoeba. The first cyclic sterol precursor is cycloartenol, which is the sterol precursor in all photosynthetic phyla. No trace of lanosterol, which is the sterol precursor in animals and fungi, could be detected. These results show that at least some of the biochemical processes of Acanthamoeba polyphaga might be phylogenetically related to those of unicellular algae. Addition of exogenous sterols to the culture medium does not influence the sterol biosynthesis and the sterol composition of the cells.
54 citations
TL;DR: Measurement of stoichiometry of 5-desaturase demonstrates that for each equivalent of cis-desaturation of delta 7-cholestenol, 1 eq of NADH is consumed, which confirms, as suggested by previous workers, that the 5- Desaturase is catalyzed by a mixed function oxidase rather than a dehydrogenase.
50 citations
TL;DR: The results obtained from this study indicate that repression of ergosterol synthesis by naphthiomate and miconazole is due to inhibition of squalene epoxidation for the former and C14-demethylation for the latter.
36 citations
TL;DR: The Noxides of several tertiary amines possessing an aliphatic long chain are more potent inhibitors of 2,3-epoxysqualene-β-amyrin and -lanosterol cyclases than their parent amines as mentioned in this paper.
Abstract: The N-oxides of several tertiary amines possessing an aliphatic long chain are more potent inhibitors of 2,3-epoxysqualene-β-amyrin and -lanosterol cyclases than their parent amines.
22 citations
TL;DR: An altered cytochrome P-450 was partially purified from Saccharomyces cerevisiae mutant SG1 which is defective in lanosterol 14 alpha-demethylation, suggesting the presence of a nitrogenous ligand such as histidine of the apoprotein at the 6th coordination position.
TL;DR: Two nystatin-resistant strains of S. cerevisiae have been compared and found to have differences in their reduced-CO difference spectra indicating different distortions in the enzyme molecule.
Abstract: A number of nystatin-resistant strains of S. cerevisiae have been isolated which are defective in lanosterol C-14 demethylation, a reaction normally catalysed by cytochrome P-450. In this paper two of these strains have been compared and found to have differences in their reduced-CO difference spectra indicating different distortions in the enzyme molecule. Nystatin resistance in the C-14 demethylation deficient SG1 in shown to be determined by a single gene, and a sterol 5,6-desaturase defect does not appear to be required for viability of SG1, was reported for the C-14 demethylase deficient isolate JR4 by Taylor et al. (1983). There are at least two discernable mutant phenotypes for the yeast cytochrome P-450 structural gene which give a C-14 demethylase defect.
TL;DR: In higher organisms, this pathway is primarily restricted to the liver, small intestine, kidney, and endocrine organs, while other classes of cells do maintain the enzymes for this set of reactions, they depend upon the liver as a source of sterol.
Abstract: Publisher Summary This chapter discusses the pathway of cholesterol biosynthesis. The pathway is divided into segments that correspond to the chemical and biochemical divisions of biosynthetic route. The initial part of the pathway is the three-step conversion of acetyl-CoA to 3-hydroxy-3-methylglutaryl-Co(HMG-CoA). The next is the reduction of this molecule to mevalonate, considered to be the rate-controlling step in the biosynthesis of polyisoprenoids. Thence, a series of phosphorylation reactions both activate and decarboxylate mevalonate to isopentenyl pyrophosphate, the true isoprenoid precursor. After a rearrangement to the allylic pyrophosphate, dimethylallyl pyrophosphate, a sequence of 1'-4 condensations between the allylic and homoallylic pyrophosphates leads to the synthesis of prenyl pyrophosphates that are requisite for the synthesis of the higher polyprenols, such as cholesterol, dolichol, and coenzyme Q. For the synthesis of cholesterol, the polymerization is halted at the sesquiterpene level. Two molecules of the farnesyl pyrophosphate thus formed condense in head-to-head producing presqualene pyrophosphate. After a reductive rearrangement of the carbon skeleton to squalene, an epoxidation leads to the formation of 2,3-oxidosqualene, which then cyclizes to lanosterol. The final stages of sterologenesis involve the removal of three methyl groups from lanosterol and the migration and reduction of double bonds to give cholesterol. In higher organisms, this pathway is primarily restricted to the liver, small intestine, kidney, and endocrine organs. While other classes of cells do maintain the enzymes for this set of reactions, they depend upon the liver as a source of sterol.
TL;DR: Cholesterol biosynthesis in cultured human skin fibroblasts was studied by [14C]acetic acid incorporation into non-saponifiable lipids and quantification of labeled cholesterol among its precursors, indicating the existence of a relationship between endogenous cholesterol synthesis and cell growth and sheds additional light on the role of post-HMG-CoA regulation in this phenomenon.
TL;DR: It is evident that the rat liver and yeast cyclases are different; the possible nature of these differences is discussed.
Abstract: 1. 1. The ability of thirteen N -(1- n -dodecyl)-heterocyclic compounds and one N -(1- n -fluoroalkyl)-heterocyclic compound to inhibit the 2,3-oxidosqualene lanosterol-cyclase of rat liver and yeast has been tested. 2. 2. Eight of the N -(1- n -dodecyl)-heterocycles inhibited the rat liver enzyme well but none inhibited the yeast enzyme at 100 μM concentrations. 3. 3. The N -(1-fluoroalkyl)-heterocycle inhibited the rat liver enzyme mildly but did not inhibit the yeast enzyme at 100 μM concentration. 4. 4. It is evident that the rat liver and yeast cyclases are different; the possible nature of these differences is discussed.
TL;DR: Observations indicate that buthiobate is an unique inhibitor for hepatic microsomal cytochrome P-450 system, which inhibits cholesterol biosynthesis effectively but causes a little effect on drug oxidations.
Abstract: Effects of buthiobate (S-butyl S'-p-tert-butylbenzyl N-3-pyridyldithiocarbonimidate), a fungicide which inhibits lanosterol 14 alpha-demethylation catalyzed by a cytochrome P-450 (P-450 14DM) of yeast, on cytochrome P-450 of rat liver microsomes were studied. Buthiobate bound to limited forms of cytochrome P-450 in the microsomes with three different Kd values, 0.38, 1.56 and 13.6 microM. Buthiobate inhibited lanosterol 14 alpha-demethylase activity of the microsomes with a 50%-inhibition concentration of 0.4 microM. This concentration was comparable to the lowest Kd of buthiobate for the microsomal cytochrome P-450 and also to the 50%-inhibition concentration of the inhibitor for lanosterol 14 alpha-demethylation by yeast P-450 14DM. Buthiobate partially inhibited 7-ethoxycoumalin O-deethylase activity of the microsomes but inhibited neither benzphetamine N-demethylation nor p-nitroanisole O-demethylation. These observations suggest that cytochrome P-450s catalyzing drug oxidations are rather insensitive to buthiobate. These observations indicate that buthiobate is an unique inhibitor for hepatic microsomal cytochrome P-450 system, which inhibits cholesterol biosynthesis effectively but causes a little effect on drug oxidations.
TL;DR: The evolution throughout embryonic development of the rate at which acetate was converted into sterols was studied in chick brain and liver, and cholesterol and desmosterol were the major sterols formed by chick embryo brain, followed by lanosterol and squalene.
TL;DR: A highly specific system present within the cell, that is, sterol carrier proteins, which is capable of binding sterols and transporting them to specific enzyme sites involved in cholesterol biosynthesis and utilization is discussed.
Abstract: Publisher Summary This chapter discusses a highly specific system present within the cell, that is, sterol carrier proteins, which is capable of binding sterols (or squalene) and transporting them to specific enzyme sites involved in cholesterol biosynthesis and utilization Sterol carrier protein 1 (SCP 1 ) participates in the microsomal conversion of squalene to lanosterol, while sterol carrier protein 2 (SCP 2 ) participates in the microsomal conversion of lanosterol to cholesterol In addition, SCP 2 also participates in key steps in the utilization of cholesterol, as well as in the intracellular transfer of cholesterol between cellular organelles SCP 2 significantly enhances 7 α -hydroxycholesterol formation by rat liver microsomes, utilizing either endogenous microsomal membrane cholesterol as substrate, or exogenously supplied cholesterol The chapter discusses the purification and characterization of SCP 1 and SCP 2 A similar kinetic approach has been used to measure the specific activity of rat liver cytosol or of protein fractions obtained during the purification The chapter discusses the participation of sterol carrier proteins in intracellular cholesterol metabolism
TL;DR: It was concluded that lanosterol could only influence platelet behaviour if it came from the plasma, and the concentration of the steroid in both platelets and plasma is ten fold less than that required to sensitise the platelets.
Abstract: It has been reported that lanosterol can sensitize isolated rat platelets to agonists such as ADP and thrombin (4). The purpose of this paper was to determine whether lanosterol had similar effects on human platelets and whether this was achieved by changes in membrane fluidity. Lanosterol did increase the sensitivity of human platelets, particularly to adrenaline and ADP at concentrations as low as 5 mg.L-1 when added from solutions in ethanol. At similar concentrations cholesterol, 4-cholesten-3-one or ethynyloestradiol had either no effect or were inhibitory. Measurement of membrane fluidity with diphenylhexatriene indicated that lanosterol did not affect membrane fluidity. Incubation of platelets with [4C]-mevalonic acid gave rise to a very small incorporation into lanosterol, squalene and farnesol. Sudden activation of the platelets did not accelerate lanosterol synthesis during or after platelet aggregation. It was concluded that lanosterol could only influence platelet behaviour if it came from the plasma. However the concentration of the steroid in both platelets and plasma is ten fold less than that required to sensitise the platelets.
01 Jan 1985
TL;DR: 5-Desaturase is resolved from 4-methyl sterol oxidase activity of cholesterol biosynthesis by chromatography on the immobilized cytochrome b, indicating that introduction of the A6-bond and oxidation of 4a-methyl groups are catalyzed by different terminal oxidases, but resolution affords enzymes of sufficient purity to carry out reconstitution experiments.
Abstract: Microsomal A7-sterol 5-desaturase of cholesterol biosynthesis is a multienzyme system which catalyzes the introduction of the A6-bond into A‘-cholestenol to form 7-dehydrocholesterol. The detergent-solubilized 5-desaturase has been purified more than 70-fold and resolved from electron carriers and other rat liver microsomal enzymes of sterol biosynthesis by chromatography on DEAE-Sephacel, CM-Sepharose, and immobilized cytochrome b,; the 5-desaturase had not been fully resolved from cytochrom b6 reductase in earlier work. A functional electron transport system for the 5-desaturase has been reconstituted by combining the purified 5-desaturase and electron carriers with egg phosphatidylcholine liposomes. Optimizations of conditions for reconsitution have been obtained; both cytochrome b, and NADH-cytochrome b, reductase serve as electron carriers. A pyridine nucleotide-dependent flavoprotein is required and the requirement can be satisfied with either purified cytochrome bK reductase or cytochrome P-450 reductase. Cyanide and ironchelators strikingly inhibit the 5-desaturase activity, thus suggesting that 5-desaturase is a metalloenzyme as are other well-characterized cytochrome bK-dependent oxidases. 5-Desaturase is resolved from 4-methyl sterol oxidase activity of cholesterol biosynthesis by chromatography on the immobilized cytochrome b,. This resolution of the two oxidases not only indicates that introduction of the A6-bond and oxidation of 4a-methyl groups are catalyzed by different terminal oxidases, but resolution affords enzymes of sufficient purity to carry out reconstitution experiments. A novel assay based on substrate-dependent increments of oxidation of a-NADH has been developed for measurement of 5-desaturase activity. Measurement of stoichiometry of 5-desaturase demonstrates that for each equivalent of cis-desaturation of A7-cholestenol, 1 eq of NADH is consumed. Along with strict dependence upon oxygen, this observation confirms, as suggested by previous workers, that the 5-desaturation is catalyzed by a mixed function oxidase rather than a dehydrogenase.