ChoG is the main inducible extracellular cholesterol oxidase of Rhodococcus sp. strain CECT3014

Heras, Laura Fernández de las; Mascaraque, Victoria; Fernández, E; Lloréns, Juana María Navarro; Perera, Julián; Drzyzga, Oliver

doi:10.1016/j.micres.2010.05.003

Cited by 36 publications

(19 citation statements)

References 45 publications

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“…Sojo et al (1997) also showed a decline in the intracellular cholesterol oxidase of R. erythropolis coupled with an increase in extracellular cholesterol oxidase, with kinetics that suggested the excretion of extracellular enzyme was due to the partial solubilization of membrane-bound cholesterol oxidase. Therefore, the results of the present study are in agreement with reports that intracellular cholesterol oxidase contributes slightly to cell growth, while extracellular cholesterol oxidase does not (Sojo et al, 1997;Fernandez de las Heras et al, 2011).…”

Section: 5supporting

confidence: 93%

“…6D. Unlike extracellular cholesterol oxidase, the membrane-bound enzyme is involved in cell metabolism of cholesterol, which contributes to a small degree to cell growth (Sojo et al, 1997;Fernandez de las Heras et al, 2011). The second stage in the decline of phytosterol concentration could be due to an increase in the release of extracellular cholesterol oxidase, w a t e r r e s e a r c h 5 8 ( 2 0 1 4 ) 7 1 e8 1 which has been reported to appear towards the end of the exponential growth phase (Sojo et al, 1997).…”

Section: 5mentioning

confidence: 99%

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Biotransformation of phytosterols under aerobic conditions

et al. 2014

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Section: 5supporting

confidence: 93%

Section: 5mentioning

confidence: 99%

Biotransformation of phytosterols under aerobic conditions

et al. 2014

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“…For the promoter growth experiments, cells were plated in minimal medium M457 plates containing the desired carbon source and incubated at 30 °C for 3 days. Cholesterol and AD were previously dissolved in methyl-β-cyclodextrin (CD) [26] and prepared as described [27]. Plasmids and bacterial strains used are listed in Additional file 1.…”

Section: Methodsmentioning

confidence: 99%

Functional differentiation of 3-ketosteroid Δ1-dehydrogenase isozymes in Rhodococcus ruber strain Chol-4

et al. 2017

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BackgroundThe Rhodococcus ruber strain Chol-4 genome contains at least three putative 3-ketosteroid Δ1-dehydrogenase ORFs (kstD1, kstD2 and kstD3) that code for flavoenzymes involved in the steroid ring degradation. The aim of this work is the functional characterization of these enzymes prior to the developing of different biotechnological applications.ResultsThe three R. ruber KstD enzymes have different substrate profiles. KstD1 shows preference for 9OHAD and testosterone, followed by progesterone, deoxy corticosterone AD and, finally, 4-BNC, corticosterone and 19OHAD. KstD2 shows maximum preference for progesterone followed by 5α-Tes, DOC, AD testosterone, 4-BNC and lastly 19OHAD, corticosterone and 9OHAD. KstD3 preference is for saturated steroid substrates (5α-Tes) followed by progesterone and DOC. A preliminary attempt to model the catalytic pocket of the KstD proteins revealed some structural differences probably related to their catalytic differences. The expression of kstD genes has been studied by RT-PCR and RT-qPCR. All the kstD genes are transcribed under all the conditions assayed, although an additional induction in cholesterol and AD could be observed for kstD1 and in cholesterol for kstD3. Co-transcription of some correlative genes could be stated. The transcription initiation signals have been searched, both in silico and in vivo. Putative promoters in the intergenic regions upstream the kstD1, kstD2 and kstD3 genes were identified and probed in an apramycin-promoter-test vector, leading to the functional evidence of those R. ruber kstD promoters.ConclusionsAt least three putative 3-ketosteroid Δ1-dehydrogenase ORFs (kstD1, kstD2 and kstD3) have been identified and functionally confirmed in R. ruber strain Chol-4. KstD1 and KstD2 display a wide range of substrate preferences regarding to well-known intermediaries of the cholesterol degradation pathway (9OHAD and AD) and other steroid compounds. KstD3 shows a narrower substrate range with a preference for saturated substrates. KstDs differences in their catalytic properties was somehow related to structural differences revealed by a preliminary structural modelling. Transcription of R. ruber kstD genes is driven from specific promoters. The three genes are constitutively transcribed, although an additional induction is observed in kstD1 and kstD3. These enzymes have a wide versatility and allow a fine tuning-up of the KstD cellular activity.Electronic supplementary materialThe online version of this article (doi:10.1186/s12934-017-0657-1) contains supplementary material, which is available to authorized users.

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“…Cholesterol oxidase indicator plates were prepared as previously described by Fernández de las Heras et al (10) by using agar and minimal medium with cholesterol dissolved in CD; plates contained 0.1 mg/ml odianisidine and 1 U/ml peroxidase. Extracellular cholesterol oxidase activity is indicated by the production of brown color in the agar medium around the colonies.…”

Section: Methodsmentioning

confidence: 99%

Cholesterol Degradation by Gordonia cholesterolivorans

Drzyzga

Heras

Morales

et al. 2011

Appl Environ Microbiol

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This paper reports physiological and genetic data about the type strain Gordonia cholesterolivorans, a strain that is able to degrade steroid compounds containing a long carbon side chain such as cholesterol (C 27 ), cholestenone (C 27 ), ergosterol (C 28 ), and stigmasterol (C 29 ). The length of the carbon side chain appears to be of great importance for this bacterium, as the strain is unable to grow using steroids with a shorter or nonaliphatic carbon side chain such as cholic acid (C 24 ), progesterone (C 21 ), testosterone, androsterone, 4-androstene-3,17-dione (all C 19 ), and further steroids. This study also demonstrates that the degradation of cholesterol is a quite common feature of the genus Gordonia by comparing Gordonia cholesterolivorans with some other species of this genus (e.g., G. sihwensis, G. hydrophobica, G. australis, and G. neofelifaecis). Pyrosequencing of the genome of G. cholesterolivorans led to the identification of two conventional cholesterol oxidase genes on an 8-kb and a 12.8-kb genomic fragment with genetic organizations that are quite unique as compared to the genomes of other cholesterol-degrading bacteria sequenced so far. The identified two putative cholesterol oxidases of G. cholesterolivorans are both intracellularly acting enzymes of the class I type. Whereas one of these two cholesterol oxidases (ChoOx-1) shows high identity with an oxidoreductase of the opportunistic pathogen G. bronchialis and is not transcribed during growth with cholesterol, the other one (ChoOx-2) appears phylogenetically closer to cholesterol oxidases from members of the genus Rhodococcus and is transcribed constitutively. By using targeted gene disruption, a G. cholesterolivorans ChoOx-2 gene mutant strain that was unable to grow with steroids was obtained.Gordoniae appear to be widely distributed in nature, and strains have been isolated from environments such as soil, wastewater, estuary sand, mangrove rhizosphere, oil-producing wells, sewage sludge, and activated sludge foam (1, 8), as well as from clinical samples (1, 2). The isolation of strains of the genus Gordonia with special metabolic abilities has increased the potential for its application to biodegradation and bioremediation (1). Some isolates are able to partially or totally degrade xenobiotic contaminants or macromolecules, such as rubber, (di)benzothiophene, 3-ethyl-and 3-methylpyridine, and alkanes (17, 20, 21, 22). Further studies expanded the metabolic potential of the genus Gordonia, as some isolated strains metabolize butyl benzyl phthalates (e.g., Gordonia sp. strain MTCC 4818) (6) and even hazardous nitro compounds like the explosive RDX (also known as hexogen [hexahydro-1,3,5-trinitro-1,3,5-triazine]), which are known to be recalcitrant to bacterial degradation (e.g., Gordonia sp. strain KTR9) (11, 31). All of these data show the richness of metabolic activities of gordoniae and widen our view about the possible environmental and industrial application of these bacteria.The ability to degrade steroid compounds such as chol...

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ChoG is the main inducible extracellular cholesterol oxidase of Rhodococcus sp. strain CECT3014

Cited by 36 publications

References 45 publications

Biotransformation of phytosterols under aerobic conditions

Biotransformation of phytosterols under aerobic conditions

Functional differentiation of 3-ketosteroid Δ1-dehydrogenase isozymes in Rhodococcus ruber strain Chol-4

Cholesterol Degradation by Gordonia cholesterolivorans

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