Characterization of two-step deglycosylation via oxidation by glycoside oxidoreductase and defining their subfamily

Kim, Eun‐Mi; Seo, Joo‐Hyun; Baek, Kiheon; Kim, Byung‐Gee

doi:10.1038/srep10877

Cited by 12 publications

(11 citation statements)

References 32 publications

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“…The final step for C-glycosyl bond cleavage was a ␤-elimination-like reaction to produce daidzein and compound 1. Compound 1 has been reported previously as a spontaneous decomposition product of the ␤-elimination of 3-ketocarbohydrates, such as 3-ketosucrose (25), oxidized ginsenoside compound K (26), and 3-keto-levoglucosan (23), under alkaline conditions. Similar ␤-elimination-like cleavage has been observed in glycoside hydrolase families 4 and 109 (27)(28)(29).…”

Section: Discussionmentioning

confidence: 98%

Deglycosylation of the Isoflavone C -Glucoside Puerarin by a Combination of Two Recombinant Bacterial Enzymes and 3-Oxo-Glucose

Nakamura

Zhu

Komatsu

et al. 2020

Appl Environ Microbiol

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ABSTRACT A human intestinal bacterium strain related to Dorea species, PUE, can metabolize the isoflavone C-glucoside puerarin (daidzein 8-C-glucoside) to daidzein and glucose. We reported previously that 3″-oxo-puerarin is an essential reaction intermediate in enzymatic puerarin degradation, and we characterized a bacterial enzyme, the DgpB-DgpC complex, that cleaved the C-glycosidic bond in 3″-oxo-puerarin. However, the exact enzyme catalyzing the oxidation of the C-3″ hydroxyl in puerarin has not been identified. In this study, we demonstrated that recombinant DgpA, a Gfo/Idh/MocA family oxidoreductase, catalyzed puerarin oxidation in the presence of 3-oxo-glucose as the hydride acceptor. In the redox reaction, NAD(H) functioned as the cofactor, which bound tightly but noncovalently to DgpA. Kinetics analysis of DgpA revealed that the reaction proceeded via a ping-pong mechanism. Enzymatic C-deglycosylation of puerarin was achieved by a combination of recombinant DgpA, the DgpB-DgpC complex, and 3-oxo-glucose. In addition, the metabolite derived from the sugar moiety in the 3″-oxo-puerarin-cleaving reaction catalyzed by the DgpB-DgpC complex was characterized as 1,5-anhydro-d-erythro-hex-1-en-3-ulose, suggesting that the C-glycosidic linkage is cleaved through a β-elimination-like mechanism. IMPORTANCE One important role of the gut microbiota is to metabolize dietary nutrients and supplements such as flavonoid glycosides. Ingested glycosides are metabolized by intestinal bacteria to more-absorbable aglycones and further degradation products that show beneficial effects in humans. Although numerous glycoside hydrolases that catalyze O-deglycosylation have been reported, enzymes responsible for C-deglycosylation are still limited. In this study, we characterized enzymes involved in the C-deglycosylation of puerarin from a human intestinal bacterium, PUE. Here, we report the purification and characterization of a recombinant oxidoreductase involved in C-glucoside degradation. This study provides new insights for the elucidation of mechanisms of enzymatic C-deglycosylation.

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Section: Discussionmentioning

confidence: 98%

Deglycosylation of the Isoflavone C -Glucoside Puerarin by a Combination of Two Recombinant Bacterial Enzymes and 3-Oxo-Glucose

Nakamura

Zhu

Komatsu

et al. 2020

Appl Environ Microbiol

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“…strain GIN611 with glycoside deglycosylation activity different from that of common glycosidases (GHs). Later [65] they characterized its homologs in Stenotrophomonas maltophilia, Sphingobacterium multivorum, and Agrobacterium tumefaciens strains, catalyzed the deglycosylation via the same mechanism, and suggested these enzymes as a new GMC oxidoreductase subfamily-FAD-dependent glycoside oxidoreductase (FAD-GO). Interestingly, the authors showed broad glycone and aglycon specificities for these enzymes that makes them very attractive in their industrial applications.…”

Section: Auxiliary Activity Family 3 Enzymesmentioning

confidence: 99%

“…This gives us the opportunity to suggest that Zobellia AA3 enzymes could have the same glycoside oxidase activity, identification of which may be the subject for further research. Due to the broad substrate specificity, putative Zobellia FAD-GOs are of particular interest and can be considered as promising biocatalysts for glycoside deglycosylation in food and pharmaceutical industries [65].…”

Section: Auxiliary Activity Family 3 Enzymesmentioning

confidence: 99%

Comparative Genomics and CAZyme Genome Repertoires of Marine Zobellia amurskyensis KMM 3526T and Zobellia laminariae KMM 3676T

et al. 2019

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We obtained two novel draft genomes of type Zobellia strains with estimated genome sizes of 5.14 Mb for Z. amurskyensis KMM 3526Т and 5.16 Mb for Z. laminariae KMM 3676Т. Comparative genomic analysis has been carried out between obtained and known genomes of Zobellia representatives. The pan-genome of Zobellia genus is composed of 4853 orthologous clusters and the core genome was estimated at 2963 clusters. The genus CAZome was represented by 775 GHs classified into 62 families, 297 GTs of 16 families, 100 PLs of 13 families, 112 CEs of 13 families, 186 CBMs of 18 families and 42 AAs of six families. A closer inspection of the carbohydrate-active enzyme (CAZyme) genomic repertoires revealed members of new putative subfamilies of GH16 and GH117, which can be biotechnologically promising for production of oligosaccharides and rare monomers with different bioactivities. We analyzed AA3s, among them putative FAD-dependent glycoside oxidoreductases (FAD-GOs) being of particular interest as promising biocatalysts for glycoside deglycosylation in food and pharmaceutical industries.

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“…These enzymes have not been studied biochemically in detail, yet various Rhizobium , Agrobacterium , and Stenotrophomonas species (phylum Pseudomonadota) and Deinococcus aerius (phylum Deinococcota) were shown to contain genes that are phylogenetically related to bacterial pox genes. Furthermore, the corresponding gene products were able to oxidize either various glycosides ( 25 ) or glucose ( 6 ). FAD-dependent oxidoreductases from Rhizobium , Agrobacterium , and Stenotrophomonas species oxidized the sugar moieties of a range of different ginsenoides, resulting in their deglycosylation.…”

Section: Discussionmentioning

confidence: 99%

Evolution and separation of actinobacterial pyranose and C -glycoside-3-oxidases

Kostelac,

Taborda,

Martins

et al. 2024

Appl Environ Microbiol

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FAD-dependent pyranose oxidase (POx) and C -glycoside-3-oxidase (CGOx) are both members of the glucose-methanol-choline superfamily of oxidoreductases and belong to the same sequence space. Pyranose oxidases had been studied for their oxidation of monosaccharides such as D-glucose, but recently, a bacterial C -glycoside-3-oxidase that is phylogenetically related to POx and that reacts with C -glycosides such as carminic acid, mangiferin or puerarin has been described. Since these actinobacterial CGOx enzymes belong to the same sequence space as bacterial POx, they must have evolved from the same ancestor. Here, we performed a phylogenetic analysis of actinobacterial sequences and resurrected seven ancestral enzymes of the POx/CGOx sequence space to study the evolutionary trajectory of substrate preferences for monosaccharides and C -glycosides. Clade I, with its dimeric member POx from Kitasatospora aureofaciens , shows strict preference for monosaccharides (D-glucose and D-xylose) and does not react with any of the glycosides tested. No extant member of clade II has been studied to date. The two extant members of clades III and IV, monomeric POx/CGOx from Pseudoarthrobacter siccitolerans and Streptomyces canus , oxidized both monosaccharides as well as various C -glycosides (homoorientin, isovitexin, mangiferin, and puerarin). Steady-state kinetic parameters of several clades III and IV ancestral enzymes indicate that the generalist ancestor N35 slowly evolved to present-day enzymes with a much higher preference for C -glycosides than monosaccharides. Based on structural predictions of ancestors, we hypothesize that the strict specificity of bacterial clade I POx (and also fungal POx) is the result of oligomerization, which in turn results from the evolution of protein segments that were shown to be important for oligomerization, the arm, and the head domain. IMPORTANCE C -Glycosides often form active compounds in various plants. Breakage of the C-C bond in these glycosides to release the aglycone is challenging and proceeds via a two-step reaction, the oxidation of the sugar and subsequent cleavage of the C-C bond. Recently, an enzyme from a soil bacterium, FAD-dependent C -glycoside-3-oxidase (CGOx), was shown to catalyze the initial oxidation reaction. Here, we show that CGOx belongs to the same sequence space as pyranose oxidase (POx), and that an actinobacterial ancestor of the POx/CGOx family evolved into four clades, two of which show a high preference for C -glycosides.

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Characterization of two-step deglycosylation via oxidation by glycoside oxidoreductase and defining their subfamily

Cited by 12 publications

References 32 publications

Deglycosylation of the Isoflavone C -Glucoside Puerarin by a Combination of Two Recombinant Bacterial Enzymes and 3-Oxo-Glucose

Deglycosylation of the Isoflavone C -Glucoside Puerarin by a Combination of Two Recombinant Bacterial Enzymes and 3-Oxo-Glucose

Comparative Genomics and CAZyme Genome Repertoires of Marine Zobellia amurskyensis KMM 3526T and Zobellia laminariae KMM 3676T

Evolution and separation of actinobacterial pyranose and C -glycoside-3-oxidases

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