Identification and characterization of acetyl-CoA carboxylase gene cluster in Streptomyces toxytricini

Demirev, Atanas V.; Lee, Ji Seon; Sedai, Bhishma R.; Ivanov, Iván; Nam, Doo Hyun

doi:10.1007/s12275-009-0135-5

Cited by 11 publications

(15 citation statements)

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“…In S. toxytricini, the acetylCoA carboxylase gene cluster is the pccB gene locus containing accA3, pccB, and pccE, which has been demonstrated to be involved in the secondary metabolism. Disruption of the pccB and pccE genes caused a lipstatin production reduction as much as 80%, indicating their critical role in lipstatin biosynthesis (Demirev et al, 2009(Demirev et al, , 2010. The pccB gene encodes a β subunit of ACCase functioning as carboxyltransferase in the activation of ACCase along with biotinylation mediated by Bp1 protein, which needs divalent cations as a cofactor (Demirev et al, 2011).…”

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confidence: 99%

Enhanced production of lipstatin from Streptomyces toxytricini by optimizing fermentation conditions and medium

Zhu

Wang

et al. 2014

J. Gen. Appl. Microbiol.

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This paper is concerned with optimization of fermentation conditions for lipstatin production with. Under these conditions, a maximum lipstatin of 4.208 g/ml was achieved in verification experiments in 500 ml shake flasks. Key words: divalent cations; L-leucine; lipstatin; Streptomyces toxytricini IntroductionLipstatin is an irreversible inhibitor of pancreatic lipases produced by Streptomyces toxytricini Weibel et al., 1987). Lipstatin is of considerable importance as the key intermediate for the preparation of tetrahydrolipstatin (THL, Orlistat), which is a drug designed for the prophylaxis and treatment of diseases associated with obesity. Lipstatin contains a β-lactone structure that probably accounts for the irreversible lipase inhibition by covalent modification of the serine residue of its catalytic triad (Luthra et al., 2013b).The wide clinical application of orlistat has promoted the commercial production of lipstatin (Zohrabian, 2100). Hence, investigations into the improvement of the productions of lipstatin are of commercial importance. In an attempt to improve lipstatin production, improvement of lipstatinproducing strains and fermentation processes has been carried out during the past decade.S. toxytricini strains with an increased lipstatin fermentation unit have been achieved through modification and improvement by utilizing physical and chemical mutagenesis. A high lipstatin-producing strain SIPI-HJ-80 was obtained by ultraviolet ray and microwave treatment, which showed an increased productivity by 2.5 times over the original strain SIPI-UM-5 (Huang et al., 2006). More recently, a N-methyl-N -nitro-N-nitrosoguanidine (NTG) mutated strain of S. toxytricini (derived from strain ATCC 19813) was reported to have a lipstatin production of 2.88 mg/g at 264 h of fermentation time in a shake flask (Luthra et al., 2013a).With the strain S. toxytricini ATCC 19813, 8-fold enhancement (from 0.097 g/L to 0.885 g/L) in lipstatin production in a shake flask was obtained through medium optimization. It is also indicated that soya oil, soya lecithin and soya bean flour had significant effects on lipstatin production (Luthra et al., 2012). An increased lipstatin titer of 1.980 g/L was observed with selected S. toxytricini strain NRRL 15443 (Kumar et al., 2012). Further, a maximum lipstatin production of 3.290 g/L was achieved via a full factorial medium Full PaperEnhanced production of lipstatin from Streptomyces toxytricini by optimizing fermentation conditions and medium (Received February 28, 2014; Accepted April 3, 2014) None of the authors of this manuscript has any financial or personal relationship with other people or organizations that could inappropriately influence their work.

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confidence: 99%

Enhanced production of lipstatin from Streptomyces toxytricini by optimizing fermentation conditions and medium

Zhu

Wang

et al. 2014

J. Gen. Appl. Microbiol.

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“…By sequence analysis of these two fragments, the 9 ORFs were present in 11.2 kb DNA fragment referred as accD1 gene locus, which encodes the ACCase subunits related to the primary metabolism (Demirev et al, 2009).…”

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“…The recombinant proteins were expressed by the addition of 0.5 mM isopropyl-ȕ-thiogalactoside (IPTG) at 28°C when OD600 of host cell reached around 0.8, and further cultivated for 4-6 h. After sonication, the His6-tagged AccA3, PccB, and Bpl proteins were purified from the soluble fractions by column chromatography using NTA chelating agarose CL-6B resin (Peptron Inc., Korea) (Demirev et al, 2009). The purified proteins were confirmed by 12% sodium dodecyl sulfatedenatured polyacrylamide gel electrophoresis (SDS-PAGE), and assayed according to Bradford method (1976).…”

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“…Streptavidin agarose bead slurry (50% slurry suspension) (Novagen, USA) were used for detection and exlusion of the in vivo biotinylated holo-AccA3 according to the previous report (Rybak et al, 2004;Demirev et al, 2009). This procedure is required for the separation of the non-biotinylated apo-AccA3 (Į subunit).…”

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“…To confirm the enzyme activity, the separated apo-AccA3 was in vitro biotinylated by Bpl as described in the previous report (Demirev et al, 2009). The standard reaction mixture (100 ȝl) contained 20 mM glycine-NaOH (pH 11.0), 5.5 mM MgCl2, 10 mM KCl, 3 mM ATP, 0.03 mM or 0.06 mM D-biotin, 2.5 ȝM apo-AccA3, and 60 nM Bpl.…”

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Biochemical characteristization of propionyl-coenzyme a carboxylase complex of Streptomyces toxytricini

et al. 2011

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Acyl-CoA carboxylases (ACC) are involved in important primary or secondary metabolic pathways such as fatty acid and/or polyketides synthesis. In the 62 kb fragment of pccB gene locus of Streptomyces toxytricini producing a pancreatic inhibitor lipstatin, 3 distinct subunit genes of presumable propionyl-CoA carboxylase (PCCase) complex, assumed to be one of ACC responsible for the secondary metabolism, were identified along with gene for a biotin protein ligase (Bpl). The subunits of PCCase complex were a subunit (AccA3), P subunit (PccB), and auxiliary ɛ subunit (PccE). In order to disclose the involvement of the PCCase complex in secondary metabolism, some biochemical characteristics of each subunit as well as their complex were examined. In the test of substrate specificity of the PCCase complex, it was confirmed that this complex showed much higher conversion of propionyl-CoA rather than acetyl-CoA. It implies the enzyme complex could play a main role in the production of methylmalonyl-CoA from propionyl-CoA, which is a precursor of secondary polyketide biosynthesis.

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Biotechnological and industrial applications of Streptomyces metabolites

Khushboo

Kumar

Dubey

et al. 2021

Biofuels Bioprod Bioref

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Streptomyces sp. produces clinically important bioactive compounds with anticancer, antiviral, and antibacterial properties. These compounds have important roles in various fields like agriculture, and medicine, with bio-economic benefits. Progress in genome mining has resulted in the characterization of various secondary metabolites such as lipstatin, macrolide lactones, clavulanic acid, and cholesterol oxidase. Integration of DNA sequencing with computational tools has provided information regarding the biosynthetic pathways of secondary metabolites. This information is leading to the modulation of associated metabolic pathways to expand the biotechnological and industrial applications of metabolites. The increased utilization of bioactive compounds requires sustainable production to improve the bioeconomy. In this review, the authors discuss the bioactive compounds isolated from Streptomyces and advanced approaches to improve their production.

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Identification and characterization of acetyl-CoA carboxylase gene cluster in Streptomyces toxytricini

Cited by 11 publications

References 20 publications

Enhanced production of lipstatin from Streptomyces toxytricini by optimizing fermentation conditions and medium

Enhanced production of lipstatin from Streptomyces toxytricini by optimizing fermentation conditions and medium

Biochemical characteristization of propionyl-coenzyme a carboxylase complex of Streptomyces toxytricini

Biotechnological and industrial applications of Streptomyces metabolites

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