Enzyme In Sterol Biosynthesis Research Articles

Interactions between sterol biosynthesis genes in embryonic development of Arabidopsis.

The sterol biosynthesis pathway of Arabidopsis produces a large set of structurally related phytosterols including sitosterol and campesterol, the latter being the precursor of the brassinosteroids (BRs). While BRs are implicated as phytohormones in post-embryonic growth, the functions of other types of steroid molecules are not clear. Characterization of the fackel (fk) mutants provided the first hint that sterols play a role in plant embryogenesis. FK encodes a sterol C-14 reductase that acts upstream of all known enzymatic steps corresponding to BR biosynthesis mutants. Here we report that genetic screens for fk-like seedling and embryonic phenotypes have identified two additional genes coding for sterol biosynthesis enzymes: CEPHALOPOD (CPH), a C-24 sterol methyl transferase, and HYDRA1 (HYD1), a sterol C-8,7 isomerase. We describe genetic interactions between cph, hyd1 and fk, and studies with 15-azasterol, an inhibitor of sterol C-14 reductase. Our experiments reveal that FK and HYD1 act sequentially, whereas CPH acts independently of these genes to produce essential sterols. Similar experiments indicate that the BR biosynthesis gene DWF1 acts independently of FK, whereas BR receptor gene BRI1 acts downstream of FK to promote post-embryonic growth. We found embryonic patterning defects in cph mutants and describe a GC-MS analysis of cph tissues which suggests that steroid molecules in addition to BRs play critical roles during plant embryogenesis. Taken together, our results imply that the sterol biosynthesis pathway is not a simple linear pathway but a complex network of enzymes that produce essential steroid molecules for plant growth and development.

ERG6 and PDR5 regulate small lipophilic drug accumulation in yeast cells via distinct mechanisms

Diagnosis and circumvention of multi-drug resistance requires an understanding of the underlying cellular mechanisms. In the model organism Saccharomyces cerevisiae, deletions of PDR5 or ERG6 increase sensitivity to many small lipophilic drugs. Pdr5p is a plasma membrane ATP-binding cassette transporter that actively exports drugs, thereby lowering their intracellular levels. The mechanism by which ERG6, an enzyme in sterol biosynthesis, affects drug accumulation is less clear. We show here that ERG6 limits the rate of passive drug diffusion across the membrane, without affecting Pdr5p-mediated drug export. Consistent with their action by distinct mechanisms, PDR5 and ERG6 effects on drug accumulation are additive.

7-Dehydrocholesterol-dependent proteolysis of HMG-CoA reductase suppresses sterol biosynthesis in a mouse model of Smith-Lemli-Opitz/RSH syndrome.

Smith-Lemli-Opitz/RSH syndrome (SLOS), a relatively common birth-defect mental-retardation syndrome, is caused by mutations in DHCR7, whose product catalyzes an obligate step in cholesterol biosynthesis, the conversion of 7-dehydrocholesterol to cholesterol. A null mutation in the murine Dhcr7 causes an identical biochemical defect to that seen in SLOS, including markedly reduced tissue cholesterol and total sterol levels, and 30- to 40-fold elevated concentrations of 7-dehydrocholesterol. Prenatal lethality was not noted, but newborn homozygotes breathed with difficulty, did not suckle, and died soon after birth with immature lungs, enlarged bladders, and, frequently, cleft palates. Despite reduced sterol concentrations in Dhcr7(-/-) mice, mRNA levels for 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, the rate-controlling enzyme for sterol biosynthesis, the LDL receptor, and SREBP-2 appeared neither elevated nor repressed. In contrast to mRNA, protein levels and activities of HMG-CoA reductase were markedly reduced. Consistent with this finding, 7-dehydrocholesterol accelerates proteolysis of HMG-CoA reductase while sparing other key proteins. These results demonstrate that in mice without Dhcr7 activity, accumulated 7-dehydrocholesterol suppresses sterol biosynthesis posttranslationally. This effect might exacerbate abnormal development in SLOS by increasing the fetal cholesterol deficiency.

Ubiquitin-Dependent 26S Proteasomal Pathway: A Role in the Degradation of Native Human Liver CYP3A4 Expressed in Saccharomyces Cerevisiae?

Cytochrome P450, CYP3A4, is the dominant human liver endoplasmic reticulum (ER) hemoprotein enzyme, responsible for the metabolism of over 60% of clinically relevant drugs. We have previously shown that mechanism-based suicide inactivation of CYP3A4 and its rat liver ER orthologs, CYPs 3A, via heme-modification of their protein moieties, results in their ubiquitin (Ub)-dependent 26S proteasomal degradation (Korsmeyer et al. (1999) Arch. Biochem. Biophys. 365, 31; Wang et al. (1999) Arch. Biochem. Biophys. 365, 45). This is not surprising given that the heme-modified CYP3A proteins are structurally damaged. To determine whether the turnover of the native enzyme similarly recruited this pathway, we heterologously expressed this protein in wild-type Saccharomyces cerevisiae and mutant strains (hrd1Δ, hrd2-1, and hrd3Δ) previously shown to be deficient in the Ub-dependent 26S proteasomal degradation of the polytopic ER protein 3-hydroxy-3-methylglutaryl-CoA reductase (isoform Hmg2p), the rate-limiting enzyme in sterol biosynthesis, as well as in strains deficient in ER-associated Ub-conjugating enzymes, Ubc6p and/or Ubc7p (Hampton et al. (1996) Mol. Biol. Cell 7, 2029; Hampton and Bhakta (1997) Proc. Natl. Acad. Sci. USA 94, 12,944). Our findings reveal that in common with the degradation of Hmg2p, that of native CYP3A4 also requires Hrd2p (a subunit of the 19S cap complex of the 26S proteasome) and Ubc7p, and to a much lesser extent Hrd3p, a component of the ER-associated Ub-ligase complex. In contrast to Hmg2p-degradation, that of native CYP3A4 does not appear to absolutely require Hrd1p, another component of the ER-associated Ub–ligase complex. Furthermore, studies in a S. cerevisiae pep4Δ strain proven to be deficient in the vacuolar degradation of carboxypeptidase Y indicated that CYP3A4 degradation is also largely independent of vacuolar (lysosomal) proteolytic function. The degradation of two other native ER proteins, Sec61p and Sec63p, normal components of the ER translocon, were also examined in parallel and found to be stabilized to some extent in HRD2- and UBC7-deficient strains. Together these findings attest to the remarkable mechanistic diversity in the normal degradation of ER proteins.

Potent anti-Trypanosoma cruzi activities of oxidosqualene cyclase inhibitors.

Trypanosoma cruzi is the protozoan agent that causes Chagas' disease, a major health problem in Latin America. Better drugs are needed to treat infected individuals. The sterol biosynthesis pathway is a potentially excellent target for drug therapy against T. cruzi. In this study, we investigated the antitrypanosomal activities of a series of compounds designed to inhibit a key enzyme in sterol biosynthesis, oxidosqualene cyclase. This enzyme converts 2,3-oxidosqualene to the tetracyclic product, lanosterol. The lead compound, N-(4E,8E)-5,9, 13-trimethyl-4,8, 12-tetradecatrien-1-ylpyridinium, is an electron-poor aromatic mimic of a monocyclized transition state or high-energy intermediate formed from oxidosqualene. This compound and 27 related compounds were tested against mammalian-stage T. cruzi, and 12 inhibited growth by 50% at concentrations below 25 nM. The lead compound was shown to cause an accumulation of oxidosqualene and decreased production of lanosterol and ergosterol, consistent with specific inhibition of the oxidosqualene cyclase. The data demonstrate potent anti-T. cruzi activity associated with inhibition of oxidosqualene cyclase.

Sterol methyltransferase 1 controls the level of cholesterol in plants.

The side chain in plant sterols can have either a methyl or ethyl addition at carbon 24 that is absent in cholesterol. The ethyl addition is the product of two sequential methyl additions. Arabidopsis contains three genes-sterol methyltransferase 1 (SMT1), SMT2, and SMT3-homologous to yeast ERG6, which is known to encode an S-adenosylmethionine-dependent C-24 SMT that catalyzes a single methyl addition. The SMT1 polypeptide is the most similar of these Arabidopsis homologs to yeast Erg6p. Moreover, expression of Arabidopsis SMT1 in erg6 restores SMT activity to the yeast mutant. The smt1 plants have pleiotropic defects: poor growth and fertility, sensitivity of the root to calcium, and a loss of proper embryo morphogenesis. smt1 has an altered sterol content: it accumulates cholesterol and has less C-24 alkylated sterols content. Escherichia coli extracts, obtained from a strain expressing the Arabidopsis SMT1 protein, can perform both the methyl and ethyl additions to appropriate sterol substrates, although with different kinetics. The fact that smt1 null mutants still produce alkylated sterols and that SMT1 can catalyze both alkylation steps shows that there is considerable overlap in the substrate specificity of enzymes in sterol biosynthesis. The availability of the SMT1 gene and mutant should permit the manipulation of phytosterol composition, which will help elucidate the role of sterols in animal nutrition.

Purification and characterization of rat sterol 14-demethylase P450 (CYP51) expressed in Escherichia coli.

Sterol 14-demethylase P450 (CYP51) is an essential enzyme for sterol biosynthesis by eukaryotes. We have cloned rat and human CYP51 cDNAs [Aoyama, Y., Noshiro, M., Gotoh, O., Imaoka, S., Funae, Y., Kurosawa, N., Horiuchi, T., and Yoshida, Y. (1996) J. Biochem. 119, 926-933]. The cloned rat CYP51 cDNA was expressed in Escherichia coli with modification of the N-terminal amino acid sequence, and the expressed protein (CYP51m) was purified to gel-electrophoretic homogenity. The spectrophotometrically determined specific content of CYP51m was 16 nmol/mg protein and the apparent molecular weight was estimated to be 53,000 on SDS-PAGE. Soret peaks of the oxidized and reduced CO-complex of CYP51m were observed at 417 and 447 nm, respectively. The purified CYP51m catalyzed the 14-demethylation of lanosterol and 24,25-dihydrolanosterol upon reconstitution with NADPH-P450 reductase purified from rat liver microsomes. The apparent K(m) and V(max) values for lanosterol were 10.5 microM and 13.9 nmol/min/nmol P450, respectively, and those for 24, 25-dihydrolanosterol were 20.0 microM and 20.0 nmol/min/nmol P450, respectively. The lanosterol demethylase activity of the reconstituted system of CYP51m was inhibited by ketoconazole, itraconazole and fluconazole with apparent IC(50) values of 0.2, 0.7, and 160 microM, respectively.

Regulation of rat hepatic cytochrome P450 expression by sterol biosynthesis inhibition: inhibitors of squalene synthase are potent inducers of CYP2B expression in primary cultured rat hepatocytes and rat liver.

The effects of treatment with squalestatin 1, a potent inhibitor of squalene synthase, the first committed enzyme of sterol biosynthesis, were examined on cytochrome P450 expression in primary cultured rat hepatocytes and rat liver. Incubation of cultured hepatocytes with squalestatin 1 caused marked accumulations (maximal elevations that were approximately 25-100% of phenobarbital-elicited increases) of CYP2B mRNA and immunoreactive protein but not of CYP1A, CYP3A, or CYP4A. Squalestatin 1 treatment increased CYP2B and 3-hydroxy-3-methylglutaryl coenzyme A reductase mRNA content in hepatocyte cultures with comparable potencies (ED50 = 5.0 and 18 nM, respectively), and significantly induced CYP2B (mRNA, immunoreactive protein, and pentoxyresorufin O-dealkylase activity) in the livers of treated rats, producing maximal increases at a dose of 25 mg/kg/day that were approximately 32-87% of phenobarbital-induced increases. Squalestatin 1 treatment induced both CYP2B1 and CYP2B2 and activated reporter gene expression in cultured hepatocytes transiently transfected with a plasmid containing approximately 2.4 kb of CYP2B1 gene 5'-flanking region or containing a previously described phenobarbital-responsive region. Coincubation of cultured hepatocytes with 25-hydroxycholesterol suppressed squalestatin 1-mediated CYP2B and 3-hydroxy-3-methylglutaryl coenzyme A mRNA induction with approximately the same potency. Treatment of cultures with SQ-34919, a structurally distinct squalene synthase inhibitor, produced the same selective CYP2B mRNA induction as did squalestatin 1. These results suggest that inhibition of hepatic sterol synthesis activates processes that culminate in increased CYP2B gene transcription.

Dual localization of squalene epoxidase, Erg1p, in yeast reflects a relationship between the endoplasmic reticulum and lipid particles.

Squalene epoxidase, encoded by the ERG1 gene in yeast, is a key enzyme of sterol biosynthesis. Analysis of subcellular fractions revealed that squalene epoxidase was present in the microsomal fraction (30,000 x g) and also cofractionated with lipid particles. A dual localization of Erg1p was confirmed by immunofluorescence microscopy. On the basis of the distribution of marker proteins, 62% of cellular Erg1p could be assigned to the endoplasmic reticulum and 38% to lipid particles in late logarithmic-phase cells. In contrast, sterol Delta24-methyltransferase (Erg6p), an enzyme catalyzing a late step in sterol biosynthesis, was found mainly in lipid particles cofractionating with triacylglycerols and steryl esters. The relative distribution of Erg1p between the endoplasmic reticulum and lipid particles changes during growth. Squalene epoxidase (Erg1p) was absent in an erg1 disruptant strain and was induced fivefold in lipid particles and in the endoplasmic reticulum when the ERG1 gene was overexpressed from a multicopy plasmid. The amount of squalene epoxidase in both compartments was also induced approximately fivefold by treatment of yeast cells with terbinafine, an inhibitor of the fungal squalene epoxidase. In contrast to the distribution of the protein, enzymatic activity of squalene epoxidase was only detectable in the endoplasmic reticulum but was absent from isolated lipid particles. When lipid particles of the wild-type strain and microsomes of an erg1 disruptant were mixed, squalene epoxidase activity was partially restored. These findings suggest that factor(s) present in the endoplasmic reticulum are required for squalene epoxidase activity. Close contact between lipid particles and endoplasmic reticulum may be necessary for a concerted action of these two compartments in sterol biosynthesis.

Synthesis and biological activity of a squalenoid maleimide and other classes of squalene derivatives as irreversible inhibitors of 2,3-oxidosqualene cyclase

New classes of squalene derivatives were rationally designed and synthesized as irreversible inhibitors of 2,3-oxido-squalene cyclase (OSC), a key enzyme in sterol biosynthesis. The derivatives synthesized were maleimide 5, disulfides 8–9, α,β-unsaturated nitriles 10–11 and oxirane 12. The inhibitor activities of these derivatives were determined in vitro on OSC associated with pig liver microsomes. Squalene and dodecyl maleimide were the best inhibitors of OSC, showing a time-dependent enzyme inactivation. Moreover 2,3-oxidosqualene (OS), the natural substrate of OSC, partially protected the enzyme from squalene maleimide inactivation whereas the dodecyl derivative did not. This fact and the complex kinetics shown by squalene maleimide suggest the presence of different classes of thiolic groups essential to the activity of OSC.

Development of improved cell-based assays and screens in Saccharomyces through the combination of molecular and classical genetics.

Traditionally, the discovery of pharmaceutical and agrochemical products has largely depended on mass screening. Over the years, screen design and screening programs have evolved in terms of the sensitivity with which active material can be identified, the number of samples that can be tested, and the types of molecular targets and cellular functions that can be conveniently assayed. More recently, screens with desirable properties have been developed for a great variety of molecular targets through the exploitation of Saccharomyces molecular biology and genetics. Recent advances have enabled researchers to develop yeast-based screens for agents acting on a number of new therapeutic targets: G-protein linked receptors, cytoplasmic receptors, ion (potassium) channels, novel fungal cell wall enzymes, fungal sterol biosynthesis enzymes, antiviral targets, immunosuppressive targets, cyclic nucleotide phosphodiesterase, oncogenes and the multiple drug resistance (MDR) protein.

Physiological roles of acetoacetyl-CoA thiolase in n-alkane-utilizable yeast, Candida tropicalis: possible contribution to alkane degradation and sterol biosynthesis.

The presence of two types of thiolases, acetoacetyl-CoA thiolase and 3-ketoacyl-CoA thiolase, was demonstrated in peroxisomes of n-alkane-grown Candida tropicalis [Kurihara, T., Ueda, M., & Tanaka, A. (1989) J. Biochem. 106, 474-478], while acetoacetyl-CoA thiolase was also shown to be present in cytosol. The activity of the enzyme in cytosol was constant irrespective of culture conditions, while the peroxisomal enzyme was inducibly synthesized in the alkane-grown yeast cells. These results indicate that peroxisomal acetoacetyl-CoA thiolase participates in alkane degradation, while the cytosolic enzyme is associated with other fundamental metabolic processes, probably sterol biosynthesis, because this enzyme can catalyze the first step of the sterol biosynthesis. 3-Hydroxy-3-methylglutaryl (HMG)-CoA reductase, a key regulatory enzyme of sterol biosynthesis, was found to be localized exclusively in microsomes of the alkane-grown yeast cells. These results suggest that yeast peroxisomes do not contribute to sterol biosynthesis, unlike the case of mammalian cells.

Bisphosphonates used for the treatment of bone disorders inhibit squalene synthase and cholesterol biosynthesis.

Some bisphosphonates used for the treatment of bone disorders are also potent inhibitors of squalene synthase, a critical enzyme for sterol biosynthesis. Among seven drugs tested, YM 175 (cycloheptylaminomethylene-1,1-bisphosphonic acid) was the most potent inhibitor of rat liver microsomal squalene synthase (Ki = 57 nM) and sterol biosynthesis from [14C]mevalonate in rat liver homogenate (IC50 = 17 nM). EB 1053 (3-(1-pyrolidino)-1-hydroxypropylidene-1,1-bisphosphonic acid) and PHPBP (3-(1-piperidino)-1-hydroxypropylidene-1,1-bisphosphonic acid) were less potent inhibitors in both these assays. Pamidronate and alendronate were poor inhibitors of squalene synthase (IC50 > 10 microM) but were potent inhibitors of sterol biosynthesis from mevalonate (IC50 = 420 and 168 nM, respectively), suggesting that the latter two agents may have inhibited other enzymes involved in the synthesis of farnesyl pyrophosphate from mevalonate. Etidronate and clodronate were inactive in both these assays. YM 175 also inhibited sterol biosynthesis in mouse macrophage J774 cells (IC50 = 64 microM) and in rats, when administered acutely, it inhibited cholesterol biosynthesis in the liver (ED50 = 30 mg/kg, s.c.). Structural modifications on YM 175 to enhance cell permeability may result in a new class of cholesterol-lowering agents.

Solubilization, Partial Purification, and Immunodetection of Squalene Synthetase from Tobacco Cell Suspension Cultures

Squalene synthetase, an integral membrane protein and the first committed enzyme for sterol biosynthesis, was solubilized and partially purified from tobacco (Nicotiana tabacum) cell suspension cultures. Tobacco microsomes were prepared and the enzyme was solubilized from the lipid bilayer using a two-step procedure. Microsomes were initially treated with concentrations of octyl-beta-d-thioglucopyranoside and glycodeoxycholate below their critical micelle concentration, 4.5 and 1.1 millimolar, respectively, to remove loosely associated proteins. Complete solubilization of the squalene synthetase enzyme activity was achieved after a second treatment at detergent concentrations above or at their critical micelle concentration, 18 and 2.2 millimolar, respectively. The detergent-solubilized enzyme was further purified by a combination of ultrafiltration, gel permeation, and Fast Protein Liquid Chromatography anion exchange. A 60-fold purification and 20% recovery of the enzyme activity was achieved. The partially purified squalene synthetase protein was used to generate polyclonal antibodies from mice that efficiently inhibited synthetase activity in an in vitro assay. The apparent molecular mass of the squalene synthetase protein as determined by immunoblot analysis of the partially purified squalene synthetase protein separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis was 47 kilodaltons. The partially purified squalene synthetase activity was optimal at pH 6.0, exhibited a K(m) for farnesyl diphosphate of 9.5 micromolar, and preferred NADPH as a reductant rather than NADH.

Inhibition of sterol biosynthesis enzymes in vitro by analogues of high-energy carbocationic intermediates

Inhibition of sterol biosynthesis enzymes <i>in vitro</i> by analogues of high-energy carbocationic intermediates

Sterol-independent Regulation of 3-Hydroxy-3-methylglutaryl-CoA Reductase by Mevalonate in Chinese Hamster Ovary Cells: Magnitude and specificity

In this paper, we assess the relative degree of regulation of the rate-limiting enzyme of isoprenoid biosynthesis, 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, by sterol and nonsterol products of mevalonate by utilizing cultured Chinese hamster ovary cells blocked in sterol synthesis. We also examine the two other enzymes of mevalonate biosynthesis, acetoacetyl-CoA thiolase and HMG-CoA synthase, for regulation by mevalonate supplements. These studies indicate that in proliferating fibroblasts, treatment with mevalonic acid can produce a suppression of HMG-CoA reductase activity similar to magnitude to that caused by oxygenated sterols. In contrast, HMG-CoA synthase and acetoacetyl-CoA thiolase are only weakly regulated by mevalonate when compared with 25-hydroxycholesterol. Furthermore, neither HMG-CoA synthase nor acetoacetyl-CoA thiolase exhibits the multivalent control response by sterol and mevalonate supplements in the absence of endogenous mevalonate synthesis which is characteristic of nonsterol regulation of HMG-CoA reductase. These observations suggest that nonsterol regulation of HMG-CoA reductase is specific to that enzyme in contrast to the pleiotropic regulation of enzymes of sterol biosynthesis observed with oxygenated sterols. In Chinese hamster ovary cells supplemented with mevalonate at concentrations that are inhibitory to reductase activity, at least 80% of the inhibition appears to be mediated by nonsterol products of mevalonate. In addition, feedback regulation of HMG-CoA reductase by endogenously synthesized nonsterol isoprenoids in the absence of exogenous sterol or mevalonate supplements also produces a 70% inhibition of the enzyme activity.

ChemInform Abstract: Synthesis of a Squalenoid Oxaziridine and Other New Classes of Squalene Derivatives, as Inhibitors of Sterol Biosynthesis.

AbstractSome new squalene derivatives such as (III)‐(V) are prepared, rationally designed as inhibitors of 2,3‐oxidosqualene (SO) cyclase, a key enzyme in sterol biosynthesis.

Synthesis of a squalenoid oxaziridine and other new classes of squalene derivatives, as inhibitors of sterol biosynthesis

New classes of squalene derivatives, rationally designed as inhibitors of 2,3-oxidosqualene (SO) cyclase, a key enzyme in sterol biosynthesis, were synthesized. These were: N-methylimine 7, aminalic hydroperoxide 8, N-methyloxaziridine 9 and N-methylamide 10.9 was synthesized by a new method, in order to prevent acid decomposition of 7 and 9. The inhibitory activities of 7–10 were determined in vitro on SO cyclase associated with rat liver and yeast microsomes, and in vivo on sterol biosynthesis in 3T3 fibroblast cultures. 9 was the best inhibitor of SO cyclase associated with rat liver and yeast microsomes. In contrast, 7 and 10 strongly inhibited biosynthesis of C 27-sterols in 3T3 fibroblast cultures.

Structural and functional conservation between yeast and human 3-hydroxy-3-methylglutaryl coenzyme A reductases, the rate-limiting enzyme of sterol biosynthesis.

The pathway of sterol biosynthesis is highly conserved in all eucaryotic cells. We demonstrated structural and functional conservation of the rate-limiting enzyme of the mammalian pathway, 3-hydroxy-3-methyl-glutaryl coenzyme A reductase (HMG-CoA reductase), between the yeast Saccharomyces cerevisiae and humans. The amino acid sequence of the two yeast HMG-CoA reductase isozymes was deduced from DNA sequence analysis of the HMG1 and HMG2 genes. Extensive sequence similarity existed between the region of the mammalian enzyme encoding the active site and the corresponding region of the two yeast isozymes. Moreover, each of the yeast isozymes, like the mammalian enzyme, contained seven potential membrane-spanning domains in the NH2-terminal region of the protein. Expression of cDNA clones encoding either hamster or human HMG-CoA reductase rescued the viability of hmg1 hmg2 yeast cells lacking this enzyme. Thus, mammalian HMG-CoA reductase can provide sufficient catalytic function to replace both yeast isozymes in vivo. The availability of yeast cells whose growth depends on human HMG-CoA reductase may provide a microbial screen to identify new drugs that can modulate cholesterol biosynthesis.

Synthesis and biological activity of azasqualenes, bis-azasqualenes and derivatives

Azasqualenes, bis-azasqualenes and derivatives, designed as inhibitors of squalene 2,3-epoxide cyclase, a key enzyme in sterol biosynthesis, were synthesized and their in vitro activities against a variety of yeasts, fungi, gram-positive and gram-negative bacteria were determined. The synthesis involves a new method of squalene degradation, together with an unusual procedure for the aminative reduction of lipophilic aldehydes. A study of the structure—activity relationship was attempted for different biological parameters: anti-bacterial and anti-fungal activities ( MIC), inhibition of mycelial growth ( GTT), surfactant activity ( CMC) and membrane perturbation activity (induction of leakage in liposomes).