Substrate Selectivities of GH78 α-L-Rhamnosidases from Human Gut Bacteria on Dietary Flavonoid Glycosides
α-L-rhamnosidases play a key role in the metabolism and biodegradation of dietary flavonoid glycosides. We have developed a novel microplate spectrophotometric method to rapidly evaluate the conversion rates and substrate selectivities of mesophilic α-L-rhamnosidases towards citrus flavanone diglyco...
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2025-02-01
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| author | Bin-Chun Li Bingbing Wu Xueting Hou Guo-Bin Ding |
| author_facet | Bin-Chun Li Bingbing Wu Xueting Hou Guo-Bin Ding |
| author_sort | Bin-Chun Li |
| collection | DOAJ |
| description | α-L-rhamnosidases play a key role in the metabolism and biodegradation of dietary flavonoid glycosides. We have developed a novel microplate spectrophotometric method to rapidly evaluate the conversion rates and substrate selectivities of mesophilic α-L-rhamnosidases towards citrus flavanone diglycosides by combining with a high-active and thermophilic β-D-glucosidase based on UV-visible spectral differences between citrus flavanone diglycosides and the corresponding aglycones under alkaline conditions. Furthermore, catalytic activities and enzyme kinetics of four α-L-rhamnosidases from human gut bacteria on various dietary flavonoid glycosides with different glycosidic bonds from various subclasses have been explored by HPLC. The α-L-rhamnosidase BtRha78A specifically removed the rhamnose group from the flavones, flavanones and flavonols diglycosides with the α-1,6 glycosidic bonds. Moreover, BtRha78A displayed higher catalytic activities on the rutinose group at 7-OH of the aglycones than at 3-OH. HFM-RhaA preferred to catalyze the flavones, flavanones and dihydrochalcones diglycosides with the α-1,2 glycosidic linkages at the 7-OH. However, this enzyme also showed high catalytic activity on the flavonol diglycoside rutin with the α-1,6 glycosidic bonds at the 3-OH. HFM-RhaC exhibited certain hydrolytic abilities towards all flavonoid diglycosides, and displayed higher activities on the flavonoid diglycosides with the α-1,6 glycosidic bonds. HFM-Rha78 weakly hydrolyzed the flavones, flavanones and dihydrochalcones diglycosides with the α-1,2 glycosidic bonds, and the flavonols diglycosides with α-1,6 glycosidic bonds. All four α-L-rhamnosidases from human gut bacteria did not exhibit catalytic activity towards the flavonoid glycosides with the α-1 glycosidic bonds. It was revealed that the α-L-rhamnosidases from human gut bacteria possessed diverse substrate selectivity on dietary flavonoid diglycosides. The structural basis for the specificity of BtRha78A on the flavonoid diglycosides with α-1,6 glycosidic bonds and the preference of HFM-RhaA on the flavonoid diglycosides with α-1,2 glycosidic bonds have been analyzed by molecular docking. |
| format | Article |
| id | doaj-art-c92de5e1602a48cc93b349167ba6bcef |
| institution | DOAJ |
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| spelling | doaj-art-c92de5e1602a48cc93b349167ba6bcef2025-08-20T02:59:15ZengMDPI AGMolecules1420-30492025-02-0130598010.3390/molecules30050980Substrate Selectivities of GH78 α-L-Rhamnosidases from Human Gut Bacteria on Dietary Flavonoid GlycosidesBin-Chun Li0Bingbing Wu1Xueting Hou2Guo-Bin Ding3Institute of Biotechnology, Key Laboratory of Chemical Biology and Molecular Engineering of Ministry of Education, Shanxi University, Taiyuan 030006, ChinaInstitute of Biotechnology, Key Laboratory of Chemical Biology and Molecular Engineering of Ministry of Education, Shanxi University, Taiyuan 030006, ChinaInstitute of Biotechnology, Key Laboratory of Chemical Biology and Molecular Engineering of Ministry of Education, Shanxi University, Taiyuan 030006, ChinaInstitute of Biotechnology, Key Laboratory of Chemical Biology and Molecular Engineering of Ministry of Education, Shanxi University, Taiyuan 030006, Chinaα-L-rhamnosidases play a key role in the metabolism and biodegradation of dietary flavonoid glycosides. We have developed a novel microplate spectrophotometric method to rapidly evaluate the conversion rates and substrate selectivities of mesophilic α-L-rhamnosidases towards citrus flavanone diglycosides by combining with a high-active and thermophilic β-D-glucosidase based on UV-visible spectral differences between citrus flavanone diglycosides and the corresponding aglycones under alkaline conditions. Furthermore, catalytic activities and enzyme kinetics of four α-L-rhamnosidases from human gut bacteria on various dietary flavonoid glycosides with different glycosidic bonds from various subclasses have been explored by HPLC. The α-L-rhamnosidase BtRha78A specifically removed the rhamnose group from the flavones, flavanones and flavonols diglycosides with the α-1,6 glycosidic bonds. Moreover, BtRha78A displayed higher catalytic activities on the rutinose group at 7-OH of the aglycones than at 3-OH. HFM-RhaA preferred to catalyze the flavones, flavanones and dihydrochalcones diglycosides with the α-1,2 glycosidic linkages at the 7-OH. However, this enzyme also showed high catalytic activity on the flavonol diglycoside rutin with the α-1,6 glycosidic bonds at the 3-OH. HFM-RhaC exhibited certain hydrolytic abilities towards all flavonoid diglycosides, and displayed higher activities on the flavonoid diglycosides with the α-1,6 glycosidic bonds. HFM-Rha78 weakly hydrolyzed the flavones, flavanones and dihydrochalcones diglycosides with the α-1,2 glycosidic bonds, and the flavonols diglycosides with α-1,6 glycosidic bonds. All four α-L-rhamnosidases from human gut bacteria did not exhibit catalytic activity towards the flavonoid glycosides with the α-1 glycosidic bonds. It was revealed that the α-L-rhamnosidases from human gut bacteria possessed diverse substrate selectivity on dietary flavonoid diglycosides. The structural basis for the specificity of BtRha78A on the flavonoid diglycosides with α-1,6 glycosidic bonds and the preference of HFM-RhaA on the flavonoid diglycosides with α-1,2 glycosidic bonds have been analyzed by molecular docking.https://www.mdpi.com/1420-3049/30/5/980substrate selectivity?-L-rhamnosidasedietary flavonoid glycosideshuman gut bacteria |
| spellingShingle | Bin-Chun Li Bingbing Wu Xueting Hou Guo-Bin Ding Substrate Selectivities of GH78 α-L-Rhamnosidases from Human Gut Bacteria on Dietary Flavonoid Glycosides Molecules substrate selectivity ?-L-rhamnosidase dietary flavonoid glycosides human gut bacteria |
| title | Substrate Selectivities of GH78 α-L-Rhamnosidases from Human Gut Bacteria on Dietary Flavonoid Glycosides |
| title_full | Substrate Selectivities of GH78 α-L-Rhamnosidases from Human Gut Bacteria on Dietary Flavonoid Glycosides |
| title_fullStr | Substrate Selectivities of GH78 α-L-Rhamnosidases from Human Gut Bacteria on Dietary Flavonoid Glycosides |
| title_full_unstemmed | Substrate Selectivities of GH78 α-L-Rhamnosidases from Human Gut Bacteria on Dietary Flavonoid Glycosides |
| title_short | Substrate Selectivities of GH78 α-L-Rhamnosidases from Human Gut Bacteria on Dietary Flavonoid Glycosides |
| title_sort | substrate selectivities of gh78 α l rhamnosidases from human gut bacteria on dietary flavonoid glycosides |
| topic | substrate selectivity ?-L-rhamnosidase dietary flavonoid glycosides human gut bacteria |
| url | https://www.mdpi.com/1420-3049/30/5/980 |
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