2458-08-4Relevant academic research and scientific papers
Preparation and characterization of some keto-bile acid azines
Bertolasi, Valerio,Bortolini, Olga,Fantin, Giancarlo,Fogagnolo, Marco,Perrone, Daniela
, p. 756 - 764 (2007)
New acyclic dimers of ketocholanic acids with hydrazine were obtained. Crystal structure was determined for the 3,7-dihydroxy-12-ketocholanic acid azine. Some distinctive 1H NMR signals are assigned for the entire set of azines.
Search and discovery of actinobacteria capable of transforming deoxycholic and cholic acids
Deshcherevskaya, N. O.,Lobastova, T. G.,Kollerov, V. V.,Donova, M. V.,Kazantsev, A. V.
, p. S157 - S165 (2016)
The capability of 54 selected actinobacteria strains of different phyla to convert deoxycholic (DCA) and cholic (CA) acids under aerobic conditions was studied. Except for the two species, the strains did not grow on DCA (1 g/l) as a sole carbon source, but some of them effectively converted DCA performing 7β- and 9α- hydroxylation, 3α- and 12α-dehydrogenation, partial cleavage of the isoprenoic side chain and Δ4-dehydrogenation. Ursocholic acid, 9α-hydroxy-3,12-dioxo-23,24-bisnorchol-4-ene-22-oic acid, 3-keto-DCA and other end metabolites had been firstly identified in the actinobacteria strains. The total yield of 12α-hydroxy-3-oxo-chol-4-ene-24-oic and 3,12-dioxochol-4-ene-24-oic acids from DCA with Rhodococcus erythropolis VKM Ac-1152 reached 95%. Almost 80% DCA were converted to 9α-hydroxy-3,12-dioxo-23,24-bisnorchol-4-en-22-oic acid by Rhodococcus sp. MTS-77. Unlike DCA, cholic acid (CA) was confirmed to be a growth substrate for majority of the examined strains, but only three Rhodococcus strains exhibited 7α- and/or 12α-HSDH activities thus forming 7-keto-DCA and 12-keto-chenodeoxycholic acid as major products from CA. Steroid metabolites were identified by TLC, GC, MS, 1H- and 13C NMR analyses. The results may contribute to the knowledge of biocatalytic potential of diverse soil-dwelling actinobacteria towards bile acids, and could be applied at the development of novel bioprocesses for production of the valuable cholanic acids.
Preparative-Scale Regio- and Stereospecific Oxidoreduction of Cholic Acid and Dehydrocholic Acid Catalyzed by Hydroxysteroid Dehydrogenases
Riva, Sergio,Bovara, Roberto,Pasta, Piero,Carrea, Giacomo
, p. 2902 - 2906 (1986)
NAD(P)-dependent hydroxysteroid dehydrogenases were used as catalysts for the oxidoreduction of the hydroxyl-keto groups of cholic acid (3α,7α,12α-trihydroxy-5β-cholan-24-oic acid) and dehydrocholic acid (3,7,12-trioxo-5β-cholan-24-oic acid).Cholic acid was regiospecifically oxidized, on a preparative scale, at each of the three possible positions, and dehydrocholic acid regio- and stereospecifically reduced at each of the three positions.The compounds were quantitatively transformed and the products were 97-99percent pure.The assignment of product structure was made by NMR.The nicotinamide cofactors were enzymatically regenerated, in situ, with the α-ketoglutarate/glutamate dehydrogenase, formate/formate dehydrogenase or glucose/glucose dehydrogenase systems.The enzymes were employed in the free form or immobilized on Sepharose CL-4B.
Clean Enzymatic Oxidation of 12α-Hydroxysteroids to 12-Oxo-Derivatives Catalyzed by Hydroxysteroid Dehydrogenase
Tonin, Fabio,Alvarenga, Natália,Ye, Jia Zheng,Arends, Isabel W. C. E.,Hanefeld, Ulf
, p. 2448 - 2455 (2019)
The C12 specific oxidation of hydroxysteroids is an essential reaction required for the preparation of pharmaceutical ingredients like ursodeoxycholic acid (UDCA) and chenodeoxycholic acid (CDCA), which can be synthesized by Wolff-Kishner reduction of the obtained 12-oxo-hydroxysteroids. 12α-hydroxysteroid dehydrogenases (12α-HSDHs) have been shown to perform this reaction with high yields, under mild conditions and without the need of protection and deprotection steps, required in chemical synthesis. Here, the recombinant expression and biochemical characterization of the nicotinamide adenine dinucleotide (NAD+)-dependent HSDH from Eggerthella lenta (El12α-HSDH) are reported. This enzyme shows comparable properties with the well-known nicotinamide adenine dinucleotide phosphate (NADP+)-dependent enzyme from Clostridium sp. 48–50. In order to perform a viable and atom efficient enzymatic hydroxysteroid oxidation, NAD(P)H oxidase (NOX) was employed as cofactor regeneration system: NOX uses oxygen (O2) as sacrificial substrate and produces only water as side product. 10 mM of cholic acid was fully and selectively converted to 12-oxo-CDCA in 24 h. The possibility to employ this system on UCA and 7-oxo-deoxycholic acid (7-oxo-DCA) as substrates was additionally investigated. The performance of the El12α-HSDH was evaluated also in combination with a “classical” regeneration system (oxaloacetate/malate dehydrogenase) showing full conversion in 4 h. Finally, the feasibility of a catalytic aerobic-NAD+-dependent enzymatic oxidation was shown on a preparative scale (oxidation of CA to 12-oxo-CDCA) employing the El12α-HSDH-NOX system in a segmented-flow-reactor. (Figure presented.).
Anodic electrochemical oxidation of cholic acid
Medici, Alessandro,Pedrini, Paola,De Battisti, Achille,Fantin, Giancarlo,Fogagnolo, Marco,Guerrini, Alessandra
, p. 63 - 69 (2001)
Regioselectivity in the anodic electrochemical oxidation of cholic acid with different anodes is described. The oxidation with PbO2 anode affords the dehydrocholic acid in quantitative yield after 22 h. 3α,12α-Dihydroxy-7-oxo-5β-cholan-24-oic acid (59%) and 3α-hydroxy-7,12-dioxo-5β-cholan-24-oic acid (51%) are obtained stopping the reaction at lower time. The rate of the OH-oxidation is C7 > C12 > C3. The electro-oxidation with platinum foil anode gives selectively the 7-ketocholic acid in 40% yield. On the other hand, the graphite plate anode, varying the reaction conditions, produces selectively the dehydrocholic acid in quantitative yield or the 3α,12α-dihydroxy-7-oxo-5β-cholan-24-oic acid (96%) while the 3α,7α-dihydroxy-12-oxo-5β-cholan-24-oic acid (34%) is obtained together with the other oxo acids. Copyright
Efficient Synthesis of 12-Oxochenodeoxycholic Acid Using a 12α-Hydroxysteroid Dehydrogenase from Rhodococcus ruber
Shi, Shou-Cheng,You, Zhi-Neng,Zhou, Ke,Chen, Qi,Pan, Jiang,Qian, Xiao-Long,Xu, Jian-He,Li, Chun-Xiu
, p. 4661 - 4668 (2019)
12α-Hydroxysteroid dehydrogenase (12α-HSDH) has the potential to convert cheap and readily available cholic acid (CA) to 12-oxochenodeoxycholic acid (12-oxo-CDCA), a key precursor for chemoenzymatic synthesis of the therapeutic bile acid ursodeoxycholic acid (UDCA). In this work, a native nicotinamide adenine dinucleotide (NAD+)-dependent 12α-hydroxysteroid dehydrogenase (Rr12α-HSDH) from Rhodococcus ruber was identified using a structure-guided genome mining (SSGM) approach, which is based on the structure of cofactor pocket and the conserved nicotinamide cofactor binding motif alignment. Rr12α-HSDH was heterologously overexpressed in Escherichia coli BL21 (DE3), purified and characterized. The purified Rr12α-HSDH showed a high oxidative activity of 290 U mg?1protein toward CA, with a catalytic efficiency (kcat/KM) of 5.10×103 mM?1 s?1. In a preparative biotransformation (100 mL), CA (200 mM, 80 g L?1) was efficiently converted to 12-oxo-CDCA in 1 h, with a 85% isolated yield and a space-time yield (STY) of up to 1632 g L?1 d?1. Furthermore, Rr12α-HSDH was shown to be able to catalyze the oxidation of other 12α-hydroxysteroids at high substrate loads (up to 200 mM), giving the corresponding 12-oxo-hydroxysteroids in 71%–85% yields, indicating the great potential of Rr12α-HSDH as a promising biocatalyst for the synthesis of various therapeutic bile acids. (Figure presented.).
Engineering Regioselectivity of a P450 Monooxygenase Enables the Synthesis of Ursodeoxycholic Acid via 7β-Hydroxylation of Lithocholic Acid
Grobe, Sascha,Badenhorst, Christoffel P. S.,Bayer, Thomas,Hamnevik, Emil,Wu, Shuke,Grathwol, Christoph W.,Link, Andreas,Koban, Sven,Brundiek, Henrike,Gro?johann, Beatrice,Bornscheuer, Uwe T.
, p. 753 - 757 (2020/12/01)
We engineered the cytochrome P450 monooxygenase CYP107D1 (OleP) from Streptomyces antibioticus for the stereo- and regioselective 7β-hydroxylation of lithocholic acid (LCA) to yield ursodeoxycholic acid (UDCA). OleP was previously shown to hydroxylate testosterone at the 7β-position but LCA is exclusively hydroxylated at the 6β-position, forming murideoxycholic acid (MDCA). Structural and 3DM analysis, and molecular docking were used to identify amino acid residues F84, S240, and V291 as specificity-determining residues. Alanine scanning identified S240A as a UDCA-producing variant. A synthetic “small but smart” library based on these positions was screened using a colorimetric assay for UDCA. We identified a nearly perfectly regio- and stereoselective triple mutant (F84Q/S240A/V291G) that produces 10-fold higher levels of UDCA than the S240A variant. This biocatalyst opens up new possibilities for the environmentally friendly synthesis of UDCA from the biological waste product LCA.
Chenodeoxycholic acid derivative or pharmaceutically acceptable salt thereof, and preparation method and applications thereof
-
, (2020/03/12)
The invention discloses a chenodeoxycholic acid derivative with a structure as shown in general formula I or a medicinal salt thereof, and a preparation method and application of the chenodeoxycholicacid derivative. The chenodeoxycholic acid derivative can up-regulate the transcription levels of FXR mRNA and SHP mRNA, can obviously activate FXR, and can be used for preparing drugs for treating orpreventing hyperlipidemia, atherosclerosis, non-alcoholic steatohepatitis, type II diabetes mellitus and other diseases related to blood fat.
Insights into the Substrate Promiscuity of Novel Hydroxysteroid Dehydrogenases
Bertuletti, Susanna,Ferrandi, Erica Elisa,Marzorati, Stefano,Vanoni, Marta,Riva, Sergio,Monti, Daniela
, p. 2474 - 2485 (2020/05/06)
Hydroxysteroid dehydrogenases (HSDHs) are valuable biocatalysts for the regio- and stereoselective modification of steroids, bile acids and other steroid derivatives. In this work, we investigated the substrate promiscuity of this highly selective class of enzymes. In order to reach this goal, a preliminary search of HSDH homologues in in-house or public available (meta)genomes was carried out. Eight novel NAD(H)-dependent HSDHs, showing either 7α-, 7β-, or 12α-HSDH activity, and including, for the first time, enzymes from extremophilic microorganisms, were identified, recombinantly produced, and characterized. Among the novel HSDHs, four highly active (up to 92 U mg?1) NAD(H)-dependent 7β-HSDHs showing negligible similarity towards previously described 7β-HSDHs, were discovered. These enzymes, along with previously characterized HSDHs, were tested as biocatalysts for the stereoselective reduction of a panel of substrates including two α-ketoesters of pharmaceutical interest and selected ketones that partially resemble the structural features of steroids. All the reactions were coupled with a suitable cofactor regeneration system. Regarding the α-ketoesters, nearly all of the tested HSDHs showed a good activity toward the selected substrates, yielding the reduced α-hydroxyester with up to 99% conversions and enantiomeric excesses. On the other hand, only the 7β-HSDHs from Collinsella aerofaciens and Clostridium absonum showed appreciable activity toward more complex ketones, i. e., (±)-trans-1-decalone, but with interesting as well as different selectivity. (Figure presented.).
Hydroxylation of lithocholic acid by selected actinobacteria and filamentous fungi
Kollerov,Monti,Deshcherevskaya,Lobastova,Ferrandi,Larovere,Gulevskaya,Riva,Donova
, p. 370 - 378 (2013/03/28)
Selected actinobacteria and filamentous fungi of different taxonomy were screened for the ability to carry out regio- and stereospecific hydroxylation of lithocholic acid (LCA) at position 7β. The production of ursodeoxycholic acid (UDCA) was for the first time shown for the fungal strains of Bipolaris, Gibberella, Cunninghamella and Curvularia, as well as for isolated actinobacterial strains of Pseudonocardia, Saccharothrix, Amycolatopsis, Lentzea, Saccharopolyspora and Nocardia genera. Along with UDCA, chenodeoxycholic (CDCA), deoxycholic (DCA), cholic (CA), 7-ketodeoxycholic and 3-ketodeoxycholic acids were detected amongst the metabolites by some strains. A strain of Gibberella zeae VKM F-2600 expressed high level of 7β-hydroxylating activity towards LCA. Under optimized conditions, the yield of UDCA reached 90% at 1 g/L of LCA and up to 60% at a 8-fold increased substrate loading. The accumulation of the major by-product, 3-keto UDCA, was limited by using selected biotransformation media.
