Two Proteins, Mn2+, and Low Molecular Cofactor Are Required for C-Glucosyl-Cleavage of Mangiferin

Sanugul, Kanjana; Akao, Teruaki; Nakamura, Norio; Hattori, Masao

doi:10.1248/bpb.28.2035

Cited by 13 publications

(12 citation statements)

References 23 publications

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“…C‐ glucosyl‐cleaving enzyme preparations from the mangiferin‐converting Bacteroides sp. MANG (Sanugul et al ., 2005b) and from dyer's (Saito, 1990) saffron ( Carthamus tinctorius ) deglycosylating orientin have partly been characterized. However, the mechanism of C ‐glucosyl cleavage remains to be elucidated.…”

Section: Discussionmentioning

confidence: 99%

“…The inefficacy of known glucosidase inhibitors towards the cleavage of mangiferin by cell‐free extracts of Bacteroides sp. MANG indicates that the involved enzyme(s) substantially differ(s) from O ‐glycosidases (Sanugul et al ., 2005b). A free hydroxyl group adjacent to the C‐ glycosyl moiety appears to be essential for cleavage of the C ‐glycosidic bond (Che et al ., 1991; Li et al ., 2000).…”

Section: Discussionmentioning

confidence: 99%

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Deglycosylation of puerarin and other aromatic C‐glucosides by a newly isolated human intestinal bacterium

Braune

Blaut

2010

Environmental Microbiology

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The human intestinal microbiota may influence the fate of bioactive polyphenols, such as the isoflavone puerarin (daidzein 8-C-glucoside), following their oral intake. Faecal suspensions from 19 healthy subjects were tested for their ability to C-deglycosylate puerarin. Only one of these catalysed this reaction. A rod-shaped Gram-positive bacterium, strain CG19-1, capable of deglycosylating puerarin to daidzein was isolated from the corresponding suspension. However, the strictly anaerobic isolate was unable to utilize puerarin as sole carbon and energy source nor any of the tested carbohydrates. Comparative 16S rRNA gene sequence analysis indicated that strain CG19-1 is a new species of the Lachnospiraceae. Strain CG19-1 also converted other aromatic C-glucosides in addition to puerarin. The xanthone C-glucoside mangiferin was deglycosylated to norathyriol. The flavone C-glucosides homoorientin and vitexin were degraded to 3-(3,4-dihydroxyphenyl)propionic acid via luteolin and 3-(4-hydroxyphenyl)propionic acid respectively. In addition, strain CG19-1 converted flavonoid O-glucosides, but at rates that were lower than those of the C-glucosides tested. The isolate deglycosylated the isoflavone O-glucosides daidzin and genistin to daidzein and genistein respectively. Several O-glucosides of the flavones luteolin and apigenin undergoing deglycosylation were subsequently cleaved to 3-(3,4-dihydroxyphenyl)propionic acid and 3-(4-hydroxyphenyl)propionic acid respectively. Moreover, strain CG19-1 cleaved both O-desmethylangolensin and 6'-hydroxy-O-desmethylangolensin to yield 2-(4-dihydroxyphenyl)propionic acid. The corresponding cleavage product, resorcinol, was only observed for O-desmethylangolensin.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Deglycosylation of puerarin and other aromatic C‐glucosides by a newly isolated human intestinal bacterium

Braune

Blaut

2010

Environmental Microbiology

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“…[3][4][5][6][7][8][9][10][11][12] A few papers described the partial purification of C-glycosidic bond cleaving enzymes. 13,14) According to the paper of Sanugul et al, 14) one of the enzymes for Cglycosidic bond cleavage reaction of mangiferin (xanthone Cglucoside) consisted of two proteins which required Mn 2+ and undetermined low molecular cofactor(s).…”

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confidence: 99%

Enzymatic Cleavage of the C-Glucosidic Bond of Puerarin by Three Proteins, Mn2+, and Oxidized Form of Nicotinamide Adenine Dinucleotide

Nakamura

Komatsu

Hattori

et al. 2013

Biological & Pharmaceutical Bulletin

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We previously isolated the human intestinal bacterium, strain PUE, which can cleave the C-glucosidic bond of puerarin to yield its aglycone daidzein and glucose. In this study, we partially purified puerarin C-glucosidic bond cleaving enzyme from the cell-free extract of strain PUE and demonstrated that the reaction was catalyzed by at least three proteins, Mn(2+), and oxidized form of nicotinamide adenine dinucleotide (NAD(+)). We completely purified one of the proteins, called protein C, by chromatographic separation in three steps. The molecular mass of protein C was approximately 40 kDa and the amino acid sequence of its N-terminal region shows high homology to those of two putative proteins which belong to Gfo/Idh/MocA family oxidoreductase. Protein C catalyzed hydrogen-deuterium exchange reaction of puerarin to 2"-deuterated puerarin in D(2)O condition, which closely resembles those of glycoside hydrolase family 4 and 109.

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“…A pilot study indicated that a cell-free extract (CFE) of strain PUE required for manganese ion on cleaving the C-glucosyl bond of puerarin, as observed in the case of mangiferin C-glucosyl cleaving enzyme. 11) Puerarin or [6Љ,6Љ-D 2 ]puerarin was added to a CFE in the presence of MnCl 2 , and the reaction mixture was anaerobically incubated at 37°C for 6 h. The mixture was filtered through a 10 kDa membrane, and a filtrate including low molecular weight compounds was treated with a PMP reagent before analyzing by HPLC-ESI-MS. Figure 2 shows the LC-MS spectra of (a) standard PMPglucose, (b) PMP-labeled sugar from the metabolite of puerarin, and (c) PMP-labeled sugar from the metabolite of [6Љ,6Љ-D 2 ]puerarin.…”

Section: Hplc-esi-ms Analysis Of a [6؆6؆-d 2 ]Puerarin Metabolite MImentioning

confidence: 99%

“…1,[3][4][5][6][7][8][9][10][11] Despite of the growing importance of C-glycosides in the field of pharmaceuticals, characteristic features of C-glucosyl bond-cleaving enzymes involved in its metabolism were still remain to be unclear.…”

mentioning

confidence: 99%

The C-Glucosyl Bond of Puerarin Was Cleaved Hydrolytically by a Human Intestinal Bacterium Strain PUE to Yield Its Aglycone Daidzein and an Intact Glucose

Nakamura

Nishihata

Jin

et al. 2011

CHEMICAL & PHARMACEUTICAL BULLETIN

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Many types of glycosides are widely distributed as major constituents in medicinal plants. However, due to their hydrophilic properties, they are poorly absorbed from gastro intestinal tract into body fluid, but their aglycones formed by intestinal bacterial hydrolysis are readily absorbed. Therefore, intestinal bacteria play an important role in increasing absorption and efficacy of drugs including glycosides. In nature, there is a special type of glycoside, C-glycosides, such as those of flavone, isoflavone, chromone, xanthone, anthrone, and gallic acid. Since C-1 of the sugar ring is directly connected to the aglycone by C-C bonding, C-glycosides are usually resistant against acidic and enzymatic hydrolysis in contrast with the corresponding O-glycosides. Thus, with the prospect of improving the stability, various C-glycoside analogues mimicking bioactive but labile O-glycosides have been prepared for developing the new drugs.2) In addition, a lot of publications describing in detail about O-glycosidase appeared, but that of C-glycosidase did not.On the other aspects, in the last two decades, C-glucosyl bond-cleaving reactions by human intestinal bacteria were reported. 1,[3][4][5][6][7][8][9][10][11] Despite of the growing importance of C-glycosides in the field of pharmaceuticals, characteristic features of C-glucosyl bond-cleaving enzymes involved in its metabolism were still remain to be unclear.Recently, we isolated a new bacterial species named strain PUE that transforms a C-glucoside puerarin to its aglycone daidzein.1) The aglycone moiety is normally detected easily because of UV active, however, the liberated sugar segment from C-glucoside has not been yet identified. The structural elucidation of the liberated sugar moiety could yield a valuable clue in revealing the reaction mechanism (Fig. 1). In the present paper, we describe the structural elucidation of the sugar moiety after cleavage of a C-glucosyl bond of puerarin by anaerobic incubation with a human intestinal bacterium, strain PUE, and a cell-free extract. Results Synthesis of [6؆,6؆-D 2 ]puerarin (8)Since puerarin (1) has only one primary alcohol at C-6 of the sugar moiety, we targeted it for labeling with deuterium. First, 1 was treated with benzaldehyde and zinc chloride to give 2. Other hydroxyl groups of 2 were acetylated with acetic anhydride in pyridine to obtain 3. Benzylidene acetal of 3 was hydrolyzed under an acidic condition to afford 4. The primary hydroxyl of 4 was oxidized to carboxylic acid with a Jones reagent to give 5. The acetyl groups of 5 were removed by alkali to give 6, then, the carboxyl group was esterified to yield 7 by a usual method. We failed the reduction of 7 to the corresponding alcohol with LiAlD 4 due to decomposition, but 8 was prepared from 7 by slow addition of CD 3 OD into the reaction mixture containing NaBD 4 . 12) In this reaction, the addition of CH 3 OH instead of CD 3 OD gave a mixture of 8 and [6Љ-H,6Љ-D]puerarin (ca. 1 : 1 ratio), because hydrogen atoms originated from CH 3 OH were also incorp...

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Two Proteins, Mn2+, and Low Molecular Cofactor Are Required for C-Glucosyl-Cleavage of Mangiferin

Cited by 13 publications

References 23 publications

Deglycosylation of puerarin and other aromatic C‐glucosides by a newly isolated human intestinal bacterium

Deglycosylation of puerarin and other aromatic C‐glucosides by a newly isolated human intestinal bacterium

Enzymatic Cleavage of the C-Glucosidic Bond of Puerarin by Three Proteins, Mn2+, and Oxidized Form of Nicotinamide Adenine Dinucleotide

The C-Glucosyl Bond of Puerarin Was Cleaved Hydrolytically by a Human Intestinal Bacterium Strain PUE to Yield Its Aglycone Daidzein and an Intact Glucose

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