Enzymatic Biotransformation of Ginsenoside Rb1 and Gypenoside XVII into Ginsenosides Rd and F2 by Recombinant β-glucosidase from Flavobacterium johnsoniae

Hong, Hao; Cui, Chang-Hao; Kim, Jin-Kwang; Jin, Feng; Kim, Sun-Chang; Im, Wan‐Taek

doi:10.5142/jgr.2012.36.4.418

Cited by 53 publications

(37 citation statements)

References 24 publications

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“…The optimal temperature of TthBgl was 85 ºC (Fig. 2b), which is higher than the optimum for enzymes from Actinosynnema mirum (37 ºC), Azospirillum irakense (45 ºC), Flavobacterium johnsoniae (37 ºC), Martelella mediterranea (45 ºC), Myceliophthora thermophile (60 ºC), Aspergillus niger (70 ºC), and Thermoascus aurantiacus CBMAI-756 (75 ºC) (Rashid and Siddiqui 1997;Faure et al 2001;Leite et al 2008;Mao et al 2010;Hong et al 2012;Cui et al 2013;Zhao et al 2015). With good thermostability, the enzyme maintains its activity for a longer time, and the amount needed in the reaction is reduced (Pei et al 2012;Shi et al 2013).…”

Section: Biochemical and Kinetic Parametersmentioning

confidence: 97%

Cloning, Purification, and Characterization of a Thermostable β-Glucosidase from Thermotoga thermarum DSM 5069

Long¹,

Shi²,

Li³

et al. 2016

BioResources

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A 56-kDa β-glucosidase (TthBgl) derived from Thermotoga thermarum DSM 5069 was expressed and purified from Escherichia coli BL21 (DE3). The purified enzyme showed hydrolytic activity towards only pnitrophenyl-β-D-glucopyranoside among the synthetic glycosides tested. The pH maximum was 5.0, and under the conditions tested, maximal activity was at 85 ºC, and pH stability occurred from 5.0 to 6.0. After being incubated at 80 ºC for 120 min, TthBgl retained 80% of its original activity. The β-glucosidase had no apparent requirement for metal ions or other co-factors, but its activity was significantly inhibited by 0.1% SDS and 1mM Cu 2+ , in which only 3% and 10% residual activity was maintained, respectively. The Vmax of TthBgl was 8.79 U mg -1 for pnitrophenyl-β-D-glucopyranoside, while the Km was 2.41 mM. The Enzyme activity was gradually inhibited by the addition of glucose, but remained approximately 50% of its original value in 500 mM glucose. 789.25 mg/L glucose was released from cellobiose by the incubation of 0.2 U/mL TthBgl for 9 h at 75 ºC. According to a phylogenetic analysis, TthBgl belongs to the glycosyl hydrolase family 3 (GH3).

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Section: Biochemical and Kinetic Parametersmentioning

confidence: 97%

Cloning, Purification, and Characterization of a Thermostable β-Glucosidase from Thermotoga thermarum DSM 5069

Long¹,

Shi²,

Li³

et al. 2016

BioResources

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“…1). Due to the pharmacological activities of Rb1 and compound K, many researchers have sought to develop and accelerate the enzymatic biotransformation of Rb1 [7,18,25,55].…”

Section: Major Ginsenosides: Backgroundmentioning

confidence: 99%

Ginsenosides and their metabolites: a review of their pharmacological activities in the skin

et al. 2015

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Ginsenosides are representative pharmaceutical compounds found in various forms in Panax ginseng, a traditional medicinal plant. They are converted to their metabolites Rg2, Rg3, compound K, and others by human intestinal microflora following ingestion. Numerous studies have demonstrated beneficial effects of ginsenosides against aberrant molecular processes responsible for cancer, metabolic diseases and neurodegenerative diseases. Recently, antiaging effects of ginsenosides in human skin have been reported from clinical trial and in vitro model data. Ginsenosides have hence been proposed as promising natural cosmeceutical agents. In this review, we will critically review the known biological effects of several ginsenosides (Rb1, Rg3, Rd and compound K), such as anti-inflammatory and anticancer activities, which arise from the modulation of diverse molecular pathways. The application potential of ginsenosides as cosmeceutical ingredients will also be reviewed.

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“…Deglycosylated ginsenosides have been mainly obtained with glycoside hydrolase 1 (GH1) and GH3 family β-glucosidases (An et al 2010;Wang et al 2011;Hong et al 2012;Kim et al 2012;Cui et al 2013a;Cui et al 2013b;Quan et al 2013;Shin and Oh 2013;Zhao et al 2013). Among the GH3 family, β-glucosidases from Actinosynnema mirum (Cui et al 2013a), Sanguibacter keddieii , and Terrabacter ginsenosidimutans (An et al 2010) have different regioselectivity for hydrolyzing ginsenosides in spite of highly conserved active-site residues (Table 1).…”

Section: Introductionmentioning

confidence: 99%

An amino acid at position 512 in β-glucosidase from Clavibacter michiganensis determines the regioselectivity for hydrolyzing gypenoside XVII

Shin

Hong

Seo

et al. 2015

Appl Microbiol Biotechnol

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A recombinant β-glucosidase from Clavibacter michiganensis specifically hydrolyzed the outer and inner glucose linked to the C-3 position in protopanaxadiol (PPD)-type ginsenosides and the C-6 position in protopanaxatriol (PPT)-type ginsenosides except for the hydrolysis of gypenoside LXXV (GypLXXV). The enzyme converted gypenoside XVII (GypXVII) to GypLXXV by hydrolyzing the inner glucose linked to the C-3 position. The substrate-binding residues obtained from the GypXVII-docked homology models of β-glucosidase from C. michiganensis were replaced with alanine, and the amino acid residue at position 512 was selected because of the changed regioselectivity of W512A. Site-directed mutagenesis for the amino acid residue at position 512 was performed. W512A and W512K hydrolyzed the inner glucose linked to the C-3 position and the outer glucose linked to the C-20 position of GypXVII to produce GypLXXV and F2. W512R hydrolyzed only the outer glucose linked to the C-20 position of GypXVII to produce F2. However, W512E and W512D exhibited no activity for GypXVII. Thus, the amino acid at position 512 is a critical residue to determine the regioselectivity for the hydrolysis of GypXVII. These wild-type and variant enzymes produced diverse ginsenosides, including GypXVII, GypLXXV, F2, and compound K, from ginsenoside Rb1. To the best of our knowledge, this is the first report of the alteration of regioselectivity on ginsenoside hydrolysis by protein engineering.

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Enzymatic Biotransformation of Ginsenoside Rb1 and Gypenoside XVII into Ginsenosides Rd and F2 by Recombinant β-glucosidase from Flavobacterium johnsoniae

Cited by 53 publications

References 24 publications

Cloning, Purification, and Characterization of a Thermostable β-Glucosidase from Thermotoga thermarum DSM 5069

Cloning, Purification, and Characterization of a Thermostable β-Glucosidase from Thermotoga thermarum DSM 5069

Ginsenosides and their metabolites: a review of their pharmacological activities in the skin

An amino acid at position 512 in β-glucosidase from Clavibacter michiganensis determines the regioselectivity for hydrolyzing gypenoside XVII

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