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Hiroaki Yuasa - One of the best experts on this subject based on the ideXlab platform.
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Identification of the amino acid residue responsible for the Myricetin sensitivity of human proton-coupled folate transporter
Scientific Reports, 2019Co-Authors: Takahiro Yamashiro, Tomoya Yasujima, Kinya Ohta, Katsuhisa Inoue, Hiroaki YuasaAbstract:Human proton-coupled folate transporter (hPCFT/SLC46A1) has recently been found to be inhibited by Myricetin by a sustained mechanism, raising a concern that the inhibition might lead to malabsorption of folates in the intestine, where hPCFT works for their epithelial uptake. However, rat PCFT (rPCFT) has more recently been found not to be inhibited by Myricetin. Prompted by this finding, we attempted to determine the amino acid residue involved in that by analyses comparing between hPCFT and rPCFT. In the initial analysis, chimeric constructs prepared from hPCFT and rPCFT were examined for Myricetin sensitivity to determine the hPCFT segment involved in the sensitivity. Focusing on the thereby determined segment from 83rd to 186th amino acid residue, hPCFT mutants having a designated amino acid residue replaced with its counterpart in rPCFT were prepared for the subsequent analysis. Among them, only G158N-substituted hPCFT was found to be transformed to be insensitive to Myricetin and, accordingly, oppositely N158G-substituted rPCFT was transformed to be sensitive to Myricetin. These results indicate the critical role of Gly^158 in the Myricetin sensitivity of hPCFT. This finding would help advance the elucidation of the mechanism of the Myricetin-induced inhibition of hPCFT and manage the potential risk arising from that.
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Kinetic and time-dependent features of sustained inhibitory effect of Myricetin on folate transport by proton-coupled folate transporter.
Drug Metabolism and Pharmacokinetics, 2015Co-Authors: Takahiro Yamashiro, Kinya Ohta, Yayoi Hayashi, Mai Furumiya, Katsuhisa Inoue, Hiroaki YuasaAbstract:Abstract Myricetin is a flavonoid that has recently been suggested to induce sustained inhibition of proton-coupled folate transporter (PCFT/SLC46A1), which operates for intestinal folate uptake. The present study was conducted to characterize the inhibitory effect in more detail, using human PCFT stably expressed in Madin–Darby canine kidney II cells, to gain information to cope with problems potentially arising from that. The kinetics of saturable folate transport was first assessed in the absence of Myricetin in the cells pretreated with the flavonoid for 60 min. The pretreatment induced PCFT inhibition in a manner dependent on the concentration of Myricetin, where the maximum transport rate was reduced by 35.5% and 83.1%, respectively, at its concentrations of 20 μM and 50 μM. The inhibitory effect was, however, less extensive at lower folate concentrations, because the Michaelis constant was also reduced similarly in a manner dependent on Myricetin concentration. The inhibition was induced depending on the time of pretreatment and, after removal of Myricetin (50 μM) upon the manifestation of an extensive inhibition at 60 min, reversed almost completely in 90 min. This rather short time required for recovery may suggest that the sustained inhibition of PCFT is of a reversible type.
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Noncompetitive Inhibition of Proton-coupled Folate Transporter by Myricetin
Drug Metabolism and Pharmacokinetics, 2014Co-Authors: Mai Furumiya, Takahiro Yamashiro, Kinya Ohta, Yayoi Hayashi, Erina Inaoka, Katsuhisa Inoue, Chihiro Nishijima, Hiroaki YuasaAbstract:Summary: Myricetin is a flavonoid that has recently been suggested to interfere with the intestinal folate transport system. The present study was conducted to examine that possibility, focusing on its inhibitory effect on proton-coupled folate transporter (PCFT) as the molecular entity of the transport system. The uptake transport of folate was first examined in the Caco-2 cell as an intestinal epithelial cell model, and its carrier-mediated component, of which the Michaelis constant ( K m ) was 0.407 μΜ, was found to be noncompetitively inhibited by Myricetin with an inhibition constant ( K i ) of 61 μΜ. Consistent with that, folate transport by human PCFT stably expressed in Madin-Darby canine kidney II (MDCKII) cells, of which the K m was 1.246 μM, was also noncompetitively inhibited by Myricetin with a K i of 130 μM. Thus, Myricetin was suggested to inhibit intestinal folate transport by acting noncompetitively on PCFT, although the K m and K i were similarly shifted to some extent to be smaller in Caco-2 cells. Finally, epigallocatechin-3-gallate was also suggested to act in a noncompetitive manner as an inhibitory flavonoid. Care may need to be taken, therefore, in the ingestion of Myricetin and some flavonoids to maintain the absorption of folate and antifolate drugs.
Joerg Bohlmann - One of the best experts on this subject based on the ideXlab platform.
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flavonol biosynthesis genes and their use in engineering the plant antidiabetic metabolite montbretin a
Plant Physiology, 2019Co-Authors: Sandra Irmisch, Sharon Jancsik, Macaire Man Saint Yuen, Lufiani L. Madilao, Henriette Ruebsam, Joerg BohlmannAbstract:The plant metabolite montbretin A (MbA) and its precursor mini-MbA are potential new drugs for treating type 2 diabetes. These complex acylated flavonol glycosides only occur in small amounts in the corms of the ornamental plant montbretia (Crocosmia × crocosmiiflora). Our goal is to metabolically engineer Nicotiana benthamiana using montbretia genes to achieve increased production of mini-MbA and MbA. Two montbretia UDP-dependent glycosyltransferases (UGTs), CcUGT1 and CcUGT2, catalyze the formation of the first two pathway-specific intermediates in MbA biosynthesis, Myricetin 3-O-rhamnoside and Myricetin 3-O-glucosyl rhamnoside. In previous work, expression of these UGTs in N. benthamiana resulted in small amounts of kaempferol glycosides but not Myricetin glycosides, suggesting that Myricetin was limiting. Here, we investigated montbretia genes and enzymes of flavonol biosynthesis to enhance Myricetin formation in N. benthamiana. We characterized two flavanone hydroxylases, a flavonol synthase, a flavonoid 3′-hydroxylase (F3′H), and a flavonoid 3′5′-hydroxylase (F3′5′H). Montbretia flavonol synthase converted dihydroMyricetin into Myricetin. Unexpectedly, montbretia F3′5′H shared higher sequence relatedness with F3′Hs in the CYP75B subfamily of cytochromes P450 than with those with known F3′5′H activity. Transient expression of combinations of montbretia flavonol biosynthesis genes and a montbretia MYB transcription factor in N. benthamiana resulted in availability of Myricetin for MbA biosynthesis. Transient coexpression of montbretia flavonol biosynthesis genes combined with CcUGT1 and CcUGT2 in N. benthamiana resulted in 2 mg g−1 fresh weight of the MbA pathway-specific compound Myricetin 3-O-glucosyl rhamnoside. Additional expression of the montbretia acyltransferase CcAT1 led to detectable levels of mini-MbA in N. benthamiana.
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discovery of udp glycosyltransferases and bahd acyltransferases involved in the biosynthesis of the antidiabetic plant metabolite montbretin a
The Plant Cell, 2018Co-Authors: Sandra Irmisch, Sharon Jancsik, Macaire Man Saint Yuen, Lufiani L. Madilao, Christopher R Roach, Mark Oneiljohnson, Russel Williams, Stephen G Withers, Joerg BohlmannAbstract:Plant specialized metabolism serves as a rich resource of biologically active molecules for drug discovery. The acylated flavonol glycoside montbretin A (MbA) and its precursor Myricetin 3-O-(6’-O-caffeoyl)-glucosyl rhamnoside (mini-MbA) are potent inhibitors of human pancreatic α-amylase and are being developed as drug candidates to treat type-2 diabetes. MbA occurs in corms of the ornamental plant montbretia (Crocosmia x crocosmiiflora), but a system for large-scale MbA production is currently unavailable. Biosynthesis of MbA from the flavonol Myricetin and MbA accumulation occur during early stages of corm development. We established Myricetin 3-O-rhamnoside (MR), Myricetin 3-O-glucosyl rhamnoside (MRG), and mini-MbA as the first three intermediates of MbA biosynthesis. Contrasting the transcriptomes of young and old corms revealed differentially expressed UDP-sugar-dependent glycosyltransferases (UGTs) and BAHD-acyltransferases (BAHD-ATs). UGT77B2 and UGT709G2 catalyze the consecutive glycosylation of Myricetin to produce MR and of MR to give MRG, respectively. In addition, two BAHD-ATs, CcAT1 and CcAT2, catalyze the acylation of MRG to complete the formation of mini-MbA. Transcript profiles of UGT77B2, UGT709G2, CcAT1, and CcAT2 during corm development matched the metabolite profile of MbA accumulation. Expression of these enzymes in wild tobacco (Nicotiana benthamiana) resulted in the formation of a surrogate mini-MbA, validating the potential for metabolic engineering of mini-MbA in a heterologous plant system.
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discovery of udp glycosyltransferases and bahd acyltransferases involved in the biosynthesis of the antidiabetic plant metabolite montbretin a
The Plant Cell, 2018Co-Authors: Sandra Irmisch, Sharon Jancsik, Macaire Man Saint Yuen, Lufiani L. Madilao, Christopher R Roach, Mark Oneiljohnson, Russel Williams, Stephen G Withers, Seohyun Jo, Joerg BohlmannAbstract:Plant specialized metabolism serves as a rich resource of biologically active molecules for drug discovery. The acylated flavonol glycoside montbretin A (MbA) and its precursor Myricetin 3-O-(6’-O-caffeoyl)-glucosyl rhamnoside (mini-MbA) are potent inhibitors of human pancreatic α-amylase and are being developed as drug candidates to treat type-2 diabetes. MbA occurs in corms of the ornamental plant montbretia (Crocosmia x crocosmiiflora), but a system for large-scale MbA production is currently unavailable. Biosynthesis of MbA from the flavonol Myricetin and MbA accumulation occur during early stages of corm development. We established Myricetin 3-O-rhamnoside (MR), Myricetin 3-O-glucosyl rhamnoside (MRG), and mini-MbA as the first three intermediates of MbA biosynthesis. Contrasting the transcriptomes of young and old corms revealed differentially expressed UDP-sugar-dependent glycosyltransferases (UGTs) and BAHD-acyltransferases (BAHD-ATs). UGT77B2 and UGT709G2 catalyze the consecutive glycosylation of Myricetin to produce MR and of MR to give MRG, respectively. In addition, two BAHD-ATs, CcAT1 and CcAT2, catalyze the acylation of MRG to complete the formation of mini-MbA. Transcript profiles of UGT77B2, UGT709G2, CcAT1, and CcAT2 during corm development matched the metabolite profile of MbA accumulation. Expression of these enzymes in wild tobacco (Nicotiana benthamiana) resulted in the formation of a surrogate mini-MbA, validating the potential for metabolic engineering of mini-MbA in a heterologous plant system.
Takahiro Yamashiro - One of the best experts on this subject based on the ideXlab platform.
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Identification of the amino acid residue responsible for the Myricetin sensitivity of human proton-coupled folate transporter
Scientific Reports, 2019Co-Authors: Takahiro Yamashiro, Tomoya Yasujima, Kinya Ohta, Katsuhisa Inoue, Hiroaki YuasaAbstract:Human proton-coupled folate transporter (hPCFT/SLC46A1) has recently been found to be inhibited by Myricetin by a sustained mechanism, raising a concern that the inhibition might lead to malabsorption of folates in the intestine, where hPCFT works for their epithelial uptake. However, rat PCFT (rPCFT) has more recently been found not to be inhibited by Myricetin. Prompted by this finding, we attempted to determine the amino acid residue involved in that by analyses comparing between hPCFT and rPCFT. In the initial analysis, chimeric constructs prepared from hPCFT and rPCFT were examined for Myricetin sensitivity to determine the hPCFT segment involved in the sensitivity. Focusing on the thereby determined segment from 83rd to 186th amino acid residue, hPCFT mutants having a designated amino acid residue replaced with its counterpart in rPCFT were prepared for the subsequent analysis. Among them, only G158N-substituted hPCFT was found to be transformed to be insensitive to Myricetin and, accordingly, oppositely N158G-substituted rPCFT was transformed to be sensitive to Myricetin. These results indicate the critical role of Gly^158 in the Myricetin sensitivity of hPCFT. This finding would help advance the elucidation of the mechanism of the Myricetin-induced inhibition of hPCFT and manage the potential risk arising from that.
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Kinetic and time-dependent features of sustained inhibitory effect of Myricetin on folate transport by proton-coupled folate transporter.
Drug Metabolism and Pharmacokinetics, 2015Co-Authors: Takahiro Yamashiro, Kinya Ohta, Yayoi Hayashi, Mai Furumiya, Katsuhisa Inoue, Hiroaki YuasaAbstract:Abstract Myricetin is a flavonoid that has recently been suggested to induce sustained inhibition of proton-coupled folate transporter (PCFT/SLC46A1), which operates for intestinal folate uptake. The present study was conducted to characterize the inhibitory effect in more detail, using human PCFT stably expressed in Madin–Darby canine kidney II cells, to gain information to cope with problems potentially arising from that. The kinetics of saturable folate transport was first assessed in the absence of Myricetin in the cells pretreated with the flavonoid for 60 min. The pretreatment induced PCFT inhibition in a manner dependent on the concentration of Myricetin, where the maximum transport rate was reduced by 35.5% and 83.1%, respectively, at its concentrations of 20 μM and 50 μM. The inhibitory effect was, however, less extensive at lower folate concentrations, because the Michaelis constant was also reduced similarly in a manner dependent on Myricetin concentration. The inhibition was induced depending on the time of pretreatment and, after removal of Myricetin (50 μM) upon the manifestation of an extensive inhibition at 60 min, reversed almost completely in 90 min. This rather short time required for recovery may suggest that the sustained inhibition of PCFT is of a reversible type.
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Noncompetitive Inhibition of Proton-coupled Folate Transporter by Myricetin
Drug Metabolism and Pharmacokinetics, 2014Co-Authors: Mai Furumiya, Takahiro Yamashiro, Kinya Ohta, Yayoi Hayashi, Erina Inaoka, Katsuhisa Inoue, Chihiro Nishijima, Hiroaki YuasaAbstract:Summary: Myricetin is a flavonoid that has recently been suggested to interfere with the intestinal folate transport system. The present study was conducted to examine that possibility, focusing on its inhibitory effect on proton-coupled folate transporter (PCFT) as the molecular entity of the transport system. The uptake transport of folate was first examined in the Caco-2 cell as an intestinal epithelial cell model, and its carrier-mediated component, of which the Michaelis constant ( K m ) was 0.407 μΜ, was found to be noncompetitively inhibited by Myricetin with an inhibition constant ( K i ) of 61 μΜ. Consistent with that, folate transport by human PCFT stably expressed in Madin-Darby canine kidney II (MDCKII) cells, of which the K m was 1.246 μM, was also noncompetitively inhibited by Myricetin with a K i of 130 μM. Thus, Myricetin was suggested to inhibit intestinal folate transport by acting noncompetitively on PCFT, although the K m and K i were similarly shifted to some extent to be smaller in Caco-2 cells. Finally, epigallocatechin-3-gallate was also suggested to act in a noncompetitive manner as an inhibitory flavonoid. Care may need to be taken, therefore, in the ingestion of Myricetin and some flavonoids to maintain the absorption of folate and antifolate drugs.
Sandra Irmisch - One of the best experts on this subject based on the ideXlab platform.
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flavonol biosynthesis genes and their use in engineering the plant antidiabetic metabolite montbretin a
Plant Physiology, 2019Co-Authors: Sandra Irmisch, Sharon Jancsik, Macaire Man Saint Yuen, Lufiani L. Madilao, Henriette Ruebsam, Joerg BohlmannAbstract:The plant metabolite montbretin A (MbA) and its precursor mini-MbA are potential new drugs for treating type 2 diabetes. These complex acylated flavonol glycosides only occur in small amounts in the corms of the ornamental plant montbretia (Crocosmia × crocosmiiflora). Our goal is to metabolically engineer Nicotiana benthamiana using montbretia genes to achieve increased production of mini-MbA and MbA. Two montbretia UDP-dependent glycosyltransferases (UGTs), CcUGT1 and CcUGT2, catalyze the formation of the first two pathway-specific intermediates in MbA biosynthesis, Myricetin 3-O-rhamnoside and Myricetin 3-O-glucosyl rhamnoside. In previous work, expression of these UGTs in N. benthamiana resulted in small amounts of kaempferol glycosides but not Myricetin glycosides, suggesting that Myricetin was limiting. Here, we investigated montbretia genes and enzymes of flavonol biosynthesis to enhance Myricetin formation in N. benthamiana. We characterized two flavanone hydroxylases, a flavonol synthase, a flavonoid 3′-hydroxylase (F3′H), and a flavonoid 3′5′-hydroxylase (F3′5′H). Montbretia flavonol synthase converted dihydroMyricetin into Myricetin. Unexpectedly, montbretia F3′5′H shared higher sequence relatedness with F3′Hs in the CYP75B subfamily of cytochromes P450 than with those with known F3′5′H activity. Transient expression of combinations of montbretia flavonol biosynthesis genes and a montbretia MYB transcription factor in N. benthamiana resulted in availability of Myricetin for MbA biosynthesis. Transient coexpression of montbretia flavonol biosynthesis genes combined with CcUGT1 and CcUGT2 in N. benthamiana resulted in 2 mg g−1 fresh weight of the MbA pathway-specific compound Myricetin 3-O-glucosyl rhamnoside. Additional expression of the montbretia acyltransferase CcAT1 led to detectable levels of mini-MbA in N. benthamiana.
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discovery of udp glycosyltransferases and bahd acyltransferases involved in the biosynthesis of the antidiabetic plant metabolite montbretin a
The Plant Cell, 2018Co-Authors: Sandra Irmisch, Sharon Jancsik, Macaire Man Saint Yuen, Lufiani L. Madilao, Christopher R Roach, Mark Oneiljohnson, Russel Williams, Stephen G Withers, Joerg BohlmannAbstract:Plant specialized metabolism serves as a rich resource of biologically active molecules for drug discovery. The acylated flavonol glycoside montbretin A (MbA) and its precursor Myricetin 3-O-(6’-O-caffeoyl)-glucosyl rhamnoside (mini-MbA) are potent inhibitors of human pancreatic α-amylase and are being developed as drug candidates to treat type-2 diabetes. MbA occurs in corms of the ornamental plant montbretia (Crocosmia x crocosmiiflora), but a system for large-scale MbA production is currently unavailable. Biosynthesis of MbA from the flavonol Myricetin and MbA accumulation occur during early stages of corm development. We established Myricetin 3-O-rhamnoside (MR), Myricetin 3-O-glucosyl rhamnoside (MRG), and mini-MbA as the first three intermediates of MbA biosynthesis. Contrasting the transcriptomes of young and old corms revealed differentially expressed UDP-sugar-dependent glycosyltransferases (UGTs) and BAHD-acyltransferases (BAHD-ATs). UGT77B2 and UGT709G2 catalyze the consecutive glycosylation of Myricetin to produce MR and of MR to give MRG, respectively. In addition, two BAHD-ATs, CcAT1 and CcAT2, catalyze the acylation of MRG to complete the formation of mini-MbA. Transcript profiles of UGT77B2, UGT709G2, CcAT1, and CcAT2 during corm development matched the metabolite profile of MbA accumulation. Expression of these enzymes in wild tobacco (Nicotiana benthamiana) resulted in the formation of a surrogate mini-MbA, validating the potential for metabolic engineering of mini-MbA in a heterologous plant system.
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discovery of udp glycosyltransferases and bahd acyltransferases involved in the biosynthesis of the antidiabetic plant metabolite montbretin a
The Plant Cell, 2018Co-Authors: Sandra Irmisch, Sharon Jancsik, Macaire Man Saint Yuen, Lufiani L. Madilao, Christopher R Roach, Mark Oneiljohnson, Russel Williams, Stephen G Withers, Seohyun Jo, Joerg BohlmannAbstract:Plant specialized metabolism serves as a rich resource of biologically active molecules for drug discovery. The acylated flavonol glycoside montbretin A (MbA) and its precursor Myricetin 3-O-(6’-O-caffeoyl)-glucosyl rhamnoside (mini-MbA) are potent inhibitors of human pancreatic α-amylase and are being developed as drug candidates to treat type-2 diabetes. MbA occurs in corms of the ornamental plant montbretia (Crocosmia x crocosmiiflora), but a system for large-scale MbA production is currently unavailable. Biosynthesis of MbA from the flavonol Myricetin and MbA accumulation occur during early stages of corm development. We established Myricetin 3-O-rhamnoside (MR), Myricetin 3-O-glucosyl rhamnoside (MRG), and mini-MbA as the first three intermediates of MbA biosynthesis. Contrasting the transcriptomes of young and old corms revealed differentially expressed UDP-sugar-dependent glycosyltransferases (UGTs) and BAHD-acyltransferases (BAHD-ATs). UGT77B2 and UGT709G2 catalyze the consecutive glycosylation of Myricetin to produce MR and of MR to give MRG, respectively. In addition, two BAHD-ATs, CcAT1 and CcAT2, catalyze the acylation of MRG to complete the formation of mini-MbA. Transcript profiles of UGT77B2, UGT709G2, CcAT1, and CcAT2 during corm development matched the metabolite profile of MbA accumulation. Expression of these enzymes in wild tobacco (Nicotiana benthamiana) resulted in the formation of a surrogate mini-MbA, validating the potential for metabolic engineering of mini-MbA in a heterologous plant system.
Kinya Ohta - One of the best experts on this subject based on the ideXlab platform.
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Identification of the amino acid residue responsible for the Myricetin sensitivity of human proton-coupled folate transporter
Scientific Reports, 2019Co-Authors: Takahiro Yamashiro, Tomoya Yasujima, Kinya Ohta, Katsuhisa Inoue, Hiroaki YuasaAbstract:Human proton-coupled folate transporter (hPCFT/SLC46A1) has recently been found to be inhibited by Myricetin by a sustained mechanism, raising a concern that the inhibition might lead to malabsorption of folates in the intestine, where hPCFT works for their epithelial uptake. However, rat PCFT (rPCFT) has more recently been found not to be inhibited by Myricetin. Prompted by this finding, we attempted to determine the amino acid residue involved in that by analyses comparing between hPCFT and rPCFT. In the initial analysis, chimeric constructs prepared from hPCFT and rPCFT were examined for Myricetin sensitivity to determine the hPCFT segment involved in the sensitivity. Focusing on the thereby determined segment from 83rd to 186th amino acid residue, hPCFT mutants having a designated amino acid residue replaced with its counterpart in rPCFT were prepared for the subsequent analysis. Among them, only G158N-substituted hPCFT was found to be transformed to be insensitive to Myricetin and, accordingly, oppositely N158G-substituted rPCFT was transformed to be sensitive to Myricetin. These results indicate the critical role of Gly^158 in the Myricetin sensitivity of hPCFT. This finding would help advance the elucidation of the mechanism of the Myricetin-induced inhibition of hPCFT and manage the potential risk arising from that.
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Kinetic and time-dependent features of sustained inhibitory effect of Myricetin on folate transport by proton-coupled folate transporter.
Drug Metabolism and Pharmacokinetics, 2015Co-Authors: Takahiro Yamashiro, Kinya Ohta, Yayoi Hayashi, Mai Furumiya, Katsuhisa Inoue, Hiroaki YuasaAbstract:Abstract Myricetin is a flavonoid that has recently been suggested to induce sustained inhibition of proton-coupled folate transporter (PCFT/SLC46A1), which operates for intestinal folate uptake. The present study was conducted to characterize the inhibitory effect in more detail, using human PCFT stably expressed in Madin–Darby canine kidney II cells, to gain information to cope with problems potentially arising from that. The kinetics of saturable folate transport was first assessed in the absence of Myricetin in the cells pretreated with the flavonoid for 60 min. The pretreatment induced PCFT inhibition in a manner dependent on the concentration of Myricetin, where the maximum transport rate was reduced by 35.5% and 83.1%, respectively, at its concentrations of 20 μM and 50 μM. The inhibitory effect was, however, less extensive at lower folate concentrations, because the Michaelis constant was also reduced similarly in a manner dependent on Myricetin concentration. The inhibition was induced depending on the time of pretreatment and, after removal of Myricetin (50 μM) upon the manifestation of an extensive inhibition at 60 min, reversed almost completely in 90 min. This rather short time required for recovery may suggest that the sustained inhibition of PCFT is of a reversible type.
-
Noncompetitive Inhibition of Proton-coupled Folate Transporter by Myricetin
Drug Metabolism and Pharmacokinetics, 2014Co-Authors: Mai Furumiya, Takahiro Yamashiro, Kinya Ohta, Yayoi Hayashi, Erina Inaoka, Katsuhisa Inoue, Chihiro Nishijima, Hiroaki YuasaAbstract:Summary: Myricetin is a flavonoid that has recently been suggested to interfere with the intestinal folate transport system. The present study was conducted to examine that possibility, focusing on its inhibitory effect on proton-coupled folate transporter (PCFT) as the molecular entity of the transport system. The uptake transport of folate was first examined in the Caco-2 cell as an intestinal epithelial cell model, and its carrier-mediated component, of which the Michaelis constant ( K m ) was 0.407 μΜ, was found to be noncompetitively inhibited by Myricetin with an inhibition constant ( K i ) of 61 μΜ. Consistent with that, folate transport by human PCFT stably expressed in Madin-Darby canine kidney II (MDCKII) cells, of which the K m was 1.246 μM, was also noncompetitively inhibited by Myricetin with a K i of 130 μM. Thus, Myricetin was suggested to inhibit intestinal folate transport by acting noncompetitively on PCFT, although the K m and K i were similarly shifted to some extent to be smaller in Caco-2 cells. Finally, epigallocatechin-3-gallate was also suggested to act in a noncompetitive manner as an inhibitory flavonoid. Care may need to be taken, therefore, in the ingestion of Myricetin and some flavonoids to maintain the absorption of folate and antifolate drugs.
