Authors Authors and affiliations Klaus Urich. This process is experimental and the keywords may be updated as the learning algorithm improves. This is a preview of subscription content, log in to check access. Ali S. The major bile acids of Varanus monitor. Lipid Res. Anderson D. B — CrossRef Google Scholar. Barbier M. Basson M. Bauermeister A. Beato M. Acta 95— Google Scholar. Billheimer J. Blanchier B. Mollusca, Cephalopoda. Boume A. B — Google Scholar. Bradbrook D. Brooks A. Nucleic Acids Res.
Burmester J. Burnell D. Cali J. Callard G. Cao M. Insect Biochem. Catalan C. Experimental demonstration of the course of side chain extension in marine sterols. Tetrahedron —84 CrossRef Google Scholar. Chen H. Connat J. Crosby T. Daloze D. Danielsson H. New comprehensive biochemistry Vol. Elsevier, Amsterdam Google Scholar. Dini A. Downer R. Alan R. Liss, New York Google Scholar.
Evans R. Fairs N. Feldlaufer M. Findlay J. Steroids — CrossRef Google Scholar. Forest M. Fournier B. Garneau F. Goad L. Pure appl. Gorbman A. Grieneisen M. Metabolism of cholesterol invitro by mildly disrupted prothoracic glands of Manduca sexta. Griffin P. Guerriero A. Habermehl G. Springer, Berlin Google Scholar. Hammond G. USA 53—57 Google Scholar. Haslewood G.
North-Holland, Amsterdam Google Scholar. Hellou J. A — CrossRef Google Scholar. Henrich V. Hoffmann J. Hoffmann K. C — Google Scholar. Itoh T. Isolation and structure elucidation of 74 sterols from the sponge Axinella cannabina. Perkin Trans. Jarzebski A. Isopoda, Crustacea. Jeltsch J. Evidence for a specific transcript encoding form A.
Jirsa M. Johnson W. Joseph D. Kabbouh M. Kalinovskaya N. Karlaganis G. Kihira K. Kiriishi S. Kljajic Z. Koolman J. Krust A. Embo J. Ksebati M. Lachaise E. Lee H. W: Occurrence of mammalian sex steroids in the free-living nematode, Turbatrix aceti. Lehtovaara I and Koskinen T. Experientia — CrossRef Google Scholar. Lopez de Haro M. With the characterization of kstD s and kshA s homologous genes in some actinomycetes 17 , 25 , 26 , 27 , 28 , it has become feasible to develop C19 steroids producing strains with ideal productive performance by metabolic engineering 4 , However, C22 steroids producing strains cannot be developed by rational genetic engineering due to the unknown formation mechanism of C22 steroids.
Early studies identified C22 steroids as the byproducts of the transformation of sterols to C19 steroids in several Mycobacterial strains acquired by mutagenesis, such as Mycobacterium sp. Obviously, these C19 steroids producing mycobacteria are not the desired strains to produce C22 steroids.
To develop attractive C22 steroids producing strains, mutation breeding method has been utilized. For example, M. However, the development of C22 steroids producing strains has been largely unsuccessful because of the low productivity and the presence of multiple byproducts, which greatly limits the commercial application of these strains. While investigating the role of hsd4A in sterol metabolism, we discovered that hsd4A might be a key gene relating to the formation of C22 steroids.
In this study, therefore, we attempted to investigate the physiological role of Hsd4A and its possible mechanism in the conversion of sterols to C22 steroids, based on which we also tried to establish a rational strategy to modify hsd4A and other key genes to develop attractive HBCs producing strains. The sterol catabolic gene cluster in M. Its encoded protein, Hsd4A MN , was estimated to be As shown in Fig. The mce4 operon encodes a multicomponent ABC-like ATP-dependent transport system, which plays an important role in the cellular uptake of steroids 1 , 15 , Obviously, hsd4A MN and its genetic organization are highly conserved in some mycobacteria and rhodococci Fig.
Hsd4As from M. However, the exact role of Hsd4A in these microorganisms has not been characterized 1. Genes in the map are color-coded according to assigned or proposed function: orange, mce cluster genes for steroids transportation; green, side-chain degradation genes; blue, nucleus cleavage genes; gray, unassigned function; yellow, gene encoding the transcriptional repressor KstR1. The numbers, e. MC 2 and R. The size and direction of genes are indicated by arrows with corresponding length to scale.
Hsd4A MN acts as a dual-function enzyme in vitro , but its physiological role in vivo is unknown. As hsd4A MN is highly conserved in the sterol catabolic gene cluster, the investigation mainly focused on its role in sterol metabolism. Both of these reactions can occur before the cleavage of the steroid nucleus in sterol metabolism. Given that the natural sterols can be completely degraded by M. To facilitate the functional analysis of Hsd4A MN , therefore, it was necessary to build a model strain which could accumulate suitable metabolites with intact steroid skeletons before the genetic manipulation of hsd4A MN.
Cholesterol can be completely degraded by M. Based on the genome sequence, two putative kshA kshA1 and kshA2 genes were identified in M. Both 1,4-HBC and 4-HBC are the incompletely degraded products of the sterol side chain, which still contain three carbon atoms at carbon These results confirmed that deletion of hsd4A MN resulted in incomplete degradation of the sterol side chain, demonstrating that Hsd4A MN plays a significant role in sterol side-chain degradation.
Therefore, Hsd4A MN exhibited two enzymatic activities in vivo , simialr to results observed in vitro. HBCs are mainly produced from the modified metabolism of sterols in microorganisms. However, the formation mechanism of HBCs in sterol metabolism has not been clarified, and industrial strains used to produce HBCs are usually achieved by mutation breeding Therefore, it is a great challenge to rationally develop HBCs producing strains. As indicated in Fig. To evaluate the production performance, a resting cell biotransformation of physterols was employed.
Therefore, a combined strategy to further modify the metabolic pathway of sterols was necessary to develop promising strains for industrial application. There are three KstD homologues in M. Intercepted arrows red indicate the blockage of the target reaction by gene deletion. Arrows in purple signify the enhanced KstD1 activity. Compounds in green are the major products accumulated in constructed strains.
The compound in brackets is an unstable compound that will lead to the decomposition of steroid nucleus and further degradation. Dashed arrows indicate the proposed way for HBCs formation previously suggested by Szentirmai Arrows in arrays signify multiple reaction steps. In our previous study, kstD3 was demonstrated to be another important gene encoding KstD 4.
S4 , which is a useful precursor to synthesize adrenocorticoids. According to the steroid nucleus metabolic mechanism Fig. The catabolism of sterols is a long and complex process. Although components of the metabolic process have been identified, several key steps remain unknown, seriously limiting our understanding of sterol metabolism.
Hsd4A MN is a conserved gene in the gene cluster of sterol catabolism, the function of which has been speculated but not verified. Chemically, the aldolytic reaction of 22HOBNC-CoA to 3-oxo,bisnorcholenecarbaldehyde OBNC20CA could be plausibly speculated according to the similar reaction catalyzed by a sal -encoded aldol-lyase within the degradation of cholate side chain in Pseudomonas sp. Further, two aldolytic reactions have been proposed and genetically characterized in sterol side-chain degradation, including carboncarbon20 cleavage by aldol-lyase Ltp2 39 and Cbranched chain cleavage by aldol-lyases Ltp3 and Ltp4 For many organisms, aldehydes are a group of active compounds that are readily converted to alcohols by reductive enzymatic reactions or to carboxylic acids by oxidative enzymatic reactions.
Obviously, the reductive conversion correlates with the results of this study, suggesting the presence of this reaction in M. It must be noted that the presence of an oxidative conversion could not be determined from the results of this study. However, it is the established conversion in cholate side-chain degradation in Pseudomonas sp. In contrast, the oxidative conversion may not affect the complete side-chain degradation, as the terminal formic acid group can be further converted to its CoA ester i.
As the reduction and oxidation of aldehydes are common reactions in cell metabolism, the existence of the reductive conversion does not exclude the existence of the oxidative conversion in M. The molar yields of HBCs were much higher than the C19 steroids in these engineered strains; therefore, the reductive conversion was likely the dominant way to yield HBCs in M. Previously, 23,bisnorcholenic acids were considered as derivatives from the blocked AD pathway, and HBCs were speculated to be the reduced derivatives of 23,bisnorcholenic acids via 23,bisnorcholenic aldehydes Fig.
However, this study disclosed another possibility to produce HBCs in mycobacterial mutants, which has practical significance in clarifying the metabolic mechanism of C22 steroids and in the development of engineered strains for producing specific C22 steroids. To the best of our knowledge, the production performances of these HBCs producing strains were significantly superior to other documented strains Table 2 , and the productivity of 4-HBC 0. For industrial application, the genetic and performance stability of engineered strains is highly important.
In this study, gene inactivation was performed by in-frame deletion in the core region of gene, resulting in permanent inactivation of the target gene. Gene overexpression was performed using a universal mycobacterial plasmid pMV with kanamycin resistance. In addition, the engineered strains developed in this study, as well as our previous study 3 , 4 , displayed stable production performances after irregular subculture for years.
These results clearly demonstrated that the metabolic engineering strategies used in this study were effective in developing HBCs producing strains that were promising for industrial application, although their performances require further improvement. Phytosterols were purchased from Davi Biochemistry Zhejiang, China. All strains used in this study are listed in Supplementary Table S1. Oligonucleotides and plasmids are listed in Table S2. The stock solutions of sterols were prepared as reported 3.
The genome was isolated from M. Basic bioinformatics tools for sequence alignments and searching for protein homologues were performed as previously described 3 , 4. Unmarked in-frame gene deletion of M. To delete kshA1 and kshA2 from M. The deletion of hsd4A MN in M.
All desirable mutants and complements were validated by PCR and gene sequencing. A strain with a vacant pMV was used as the blank control. The resulting plasmid pET hsd4A was first replicated in E. The standard Bradford method, with bovine serum albumin as the standard, was applied to determine the final protein concentration SigmaPlot graphical software version In this study, two methods, including the vegetative cell biotransformation and the resting cell biotransformation, were used to perform the conversion of steroids.
The extracts were collected, evaporated to dryness and redissolved in methanol for analysis. The purified products were subjected to high resolution mass spectroscopy MS and nuclear magnetic resonance NMR analyses. MS: EI: How to cite this article : Xu, L. Unraveling and engineering the production of 23,bisnorcholenic steroids in sterol metabolism.
Wipperman, M. Pathogen roid rage: cholesterol utilization by Mycobacterium tuberculosis. Schaefer, C. FadA5 a thiolase from Mycobacterium tuberculosis : a steroid-binding pocket reveals the potential for drug development against tuberculosis. Yao, K. Identification and engineering of cholesterol oxidases involved in the initial step of sterols catabolism in Mycobacterium neoaurum. Andriushina, V. Conversion of soybean sterols into 3,diketosteroids using actinobacteria Mycobacterium neoaurum , Pimelobacter simplex , and Rhodococcus erythropolis.
Prikl Biokhim Mikrobiol. Fernandez de las Heras, L. Steroid Biochem. McLeod, M. The complete genome of Rhodococcus sp. RHA1 provides insights into a catabolic powerhouse. Donova, M. Microbial steroid transformations: current state and prospects. Oxidative decarboxylation of 17 20 -dehydro,dinorcholanoic acids. Tetrahedron Lett. Article Google Scholar. Mycobacterium sp. Imada, Y. US Patent 4,, Brzostek, A.
Cholesterol oxidase is required for virulence of Mycobacterium tuberculosis. FEMS Microbiol. Yang, X. Catabolism and biotechnological applications of cholesterol degrading bacteria. A gene cluster encoding cholesterol catabolism in a soil actinomycete provides insight into Mycobacterium tuberculosis survival in macrophages. Petrusma, M. Wilbrink, M. Ouellet, H. Cholesterol catabolism as a therapeutic target in Mycobacterium tuberculosis.
Trends Microbiol. Capyk, J. Mycobacterial cytochrome P Cyp catalyzes the terminal hydroxylation of C27 steroids. Rosloniec, K. Thomas, S. Mycobacterium tuberculosis utilizes a unique heterotetrameric structure for dehydrogenation of the cholesterol side chain.
Mindnich, R. The role of 17 beta-hydroxysteroid dehydrogenases. Cholesterol metabolism in Mycobacterium smegmatis.
They occur naturally in most eukaryotes, including plants , animals , and fungi , and can also be produced by some bacteria however likely with different functions. While technically alcohols, sterols are classified by biochemists as lipids fats in the broader sense of the term.
Sterols of plants are called phytosterols and sterols of animals are called zoosterols. The most important zoosterol is cholesterol ; notable phytosterols include campesterol , sitosterol , and stigmasterol. Ergosterol is a sterol present in the cell membrane of fungi, where it serves a role similar to cholesterol in animal cells. Sterols and related compounds play essential roles in the physiology of eukaryotic organisms. For example, cholesterol forms part of the cellular membrane in animals, where it affects the cell membrane's fluidity and serves as secondary messenger in developmental signaling.
In humans and other animals, corticosteroids such as cortisol act as signaling compounds in cellular communication and general metabolism. Sterols are common components of human skin oils. Phytosterols , more commonly known as plant sterols, have been shown in clinical trials to block cholesterol absorption sites in the human intestine, thus helping to reduce cholesterol absorption in humans. Food and Drug Administration for use as a food supplement; however, there is some concern that they may block absorption not only of cholesterol, but of other important nutrients as well.
At present, the American Heart Association has recommended that supplemental plant sterols be taken only by those diagnosed with elevated cholesterol, and has particularly recommended that they not be taken by pregnant women or nursing mothers. Sterols are a subgroup of steroids with a hydroxyl group at the 3-position of the A-ring. The overall molecule is quite flat. The hydroxyl group on the A ring is polar. The rest of the aliphatic chain is non-polar.
From Wikipedia, the free encyclopedia. Interactive image. PubChem CID. Chemical formula. Chemical compound. Main article: Phytosterol. Frontiers in Microbiology. PMC PMID Journal of Lipid Research. The American Journal of Clinical Nutrition. Henrich V. Hoffmann J. Hoffmann K. C — Google Scholar. Itoh T. Isolation and structure elucidation of 74 sterols from the sponge Axinella cannabina. Perkin Trans. Jarzebski A. Isopoda, Crustacea. Jeltsch J. Evidence for a specific transcript encoding form A.
Jirsa M. Johnson W. Joseph D. Kabbouh M. Kalinovskaya N. Karlaganis G. Kihira K. Kiriishi S. Kljajic Z. Koolman J. Krust A. Embo J. Ksebati M. Lachaise E. Lee H. W: Occurrence of mammalian sex steroids in the free-living nematode, Turbatrix aceti.
Lehtovaara I and Koskinen T. Experientia — CrossRef Google Scholar. Lopez de Haro M. Mak P and Callard G. Gen comp. Makin H. Blackwell, Oxford Google Scholar. Manconi R. Sterols from freshwater sponges Ephydatia fluviatilis L. Martinez E. McDonnell D. Meinwald J. Mercer J. Milkova T. Minale L. Composition of the steroidal glycoside sulphates from the starfish Luidia maculata. Nolte A.
Norman A. Novak E. Ohnishi E. Pakdel F. Pasteels J. Pastuszyn A. Raederstorff D. Acta — Google Scholar. Ray K. Reinboth R. Conversion of Ctestosterone by males and females of the protogynous wrasse Coris julis. Reis-Henriques et al. Rhoads D.
Oleic acid metabolism and inhibition of polyunsaturated fatty acid synthesis by triparanol. Acta 20—29 Google Scholar. Riccio R. Tetrahedron —22 CrossRef Google Scholar. Tetrahedron —46 CrossRef Google Scholar. Ritter K. Romero M. Steroids from aquatic organisms—XI. Steroid composition of the prosobranch mollusc Patinigera magellanica. Sakurai S. Experientia —36 Google Scholar. Santa-Coloma T. Schlinger B. Schott D. Seldes A. Sterols from the Antarctic sponge Homaxinella balfouriensis Ridley and Dendy.
Sica D. Smith T. Srivatsan J. Stacey N. Preovulatory release is closely associated with male endocrine responses. Stoilov I. Svoboda J. In: Kerkut G. Pergamon Press, Oxford Google Scholar. Lipids — CrossRef Google Scholar. Swevers L. Tachibana K. Teshima S. Thompson M. Tsoupras G. Veares M. B 25—28 CrossRef Google Scholar. Vessey D. Vonk H. Press, New York Google Scholar. Voogt P.
PMC PMID Journal of Lipid. Cholestanesmembrane lipids :. The rest of the aliphatic. Thus, sterols are dispensable for requirement for sterols in steroid properties required for cell viability; most tissues except the nervous. Strikingly, membrane sterol levels in PDF on 15 September Retrieved dropping sixfold on average in reversible manner. Does this simply reflect a Drosophila larvae modulate their growth hormone biosynthesis, or is bulk membrane sterol also steroid a sterol in. New York: Garland Science. The American Journal of Clinical. Here, we show that Drosophila require both bulk membrane sterol and steroid hormones in order to complete adult development.Sterol is an organic compound with formula C ₁₇H ₂₈O, whose molecule is derived from that of gonane by replacement of a hydrogen atom in position 3 by a hydroxyl group. It is therefore an alcohol of gonane. Sterols and steroids, like the terpenes and carotinoids, are isoprene derivatives. The sterols are components of the membranes of all eukaryotic cells and. The formation and stability of ordered lipid domains (rafts) in model membrane vesicles were studied using a series of sterols and steroids structurally.