A study on L-threonine and L-serine uptake in Escherichia coli K-12
In the current study, we report the identification and characterization of the yifK gene product as a novel amino acid carrier in E. coli K-12 cells. Both phenotypic and biochemical analyses showed that YifK acts as a permease specific to L-threonine and, to a lesser extent, L-serine. An assay of th...
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2023-03-01
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| Series: | Frontiers in Microbiology |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fmicb.2023.1151716/full |
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| author | Andrey A. Khozov Andrey A. Khozov Dmitrii M. Bubnov Eugeny D. Plisov Tatiana V. Vybornaya Tigran V. Yuzbashev Gennaro Agrimi Eugenia Messina Agnessa A. Stepanova Agnessa A. Stepanova Maxim D. Kudina Natalia V. Alekseeva Alexander I. Netrusov Sergey P. Sineoky |
| author_facet | Andrey A. Khozov Andrey A. Khozov Dmitrii M. Bubnov Eugeny D. Plisov Tatiana V. Vybornaya Tigran V. Yuzbashev Gennaro Agrimi Eugenia Messina Agnessa A. Stepanova Agnessa A. Stepanova Maxim D. Kudina Natalia V. Alekseeva Alexander I. Netrusov Sergey P. Sineoky |
| author_sort | Andrey A. Khozov |
| collection | DOAJ |
| description | In the current study, we report the identification and characterization of the yifK gene product as a novel amino acid carrier in E. coli K-12 cells. Both phenotypic and biochemical analyses showed that YifK acts as a permease specific to L-threonine and, to a lesser extent, L-serine. An assay of the effect of uncouplers and composition of the reaction medium on the transport activity indicates that YifK utilizes a proton motive force to energize substrate uptake. To identify the remaining threonine carriers, we screened a genomic library prepared from the yifK-mutant strain and found that brnQ acts as a multicopy suppressor of the threonine transport defect caused by yifK disruption. Our results indicate that BrnQ is directly involved in threonine uptake as a low-affinity but high-flux transporter, which forms the main entry point when the threonine concentration in the external environment reaches a toxic level. By abolishing YifK and BrnQ activity, we unmasked and quantified the threonine transport activity of the LIV-I branched chain amino acid transport system and demonstrated that LIV-I contributes significantly to total threonine uptake. However, this contribution is likely smaller than that of YifK. We also observed the serine transport activity of LIV-I, which was much lower compared with that of the dedicated SdaC carrier, indicating that LIV-I plays a minor role in the serine uptake. Overall, these findings allow us to propose a comprehensive model of the threonine/serine uptake subsystem in E. coli cells. |
| first_indexed | 2024-04-09T23:30:41Z |
| format | Article |
| id | doaj.art-5ea62c6237b3401d8eb7038e7f5f253a |
| institution | Directory Open Access Journal |
| issn | 1664-302X |
| language | English |
| last_indexed | 2024-04-09T23:30:41Z |
| publishDate | 2023-03-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Microbiology |
| spelling | doaj.art-5ea62c6237b3401d8eb7038e7f5f253a2023-03-21T05:38:39ZengFrontiers Media S.A.Frontiers in Microbiology1664-302X2023-03-011410.3389/fmicb.2023.11517161151716A study on L-threonine and L-serine uptake in Escherichia coli K-12Andrey A. Khozov0Andrey A. Khozov1Dmitrii M. Bubnov2Eugeny D. Plisov3Tatiana V. Vybornaya4Tigran V. Yuzbashev5Gennaro Agrimi6Eugenia Messina7Agnessa A. Stepanova8Agnessa A. Stepanova9Maxim D. Kudina10Natalia V. Alekseeva11Alexander I. Netrusov12Sergey P. Sineoky13Kurchatov Complex of Genetic Research, NRC “Kurchatov Institute”, Moscow, RussiaDepartment of Microbiology, Faculty of Biology, Lomonosov Moscow State University, Moscow, RussiaKurchatov Complex of Genetic Research, NRC “Kurchatov Institute”, Moscow, RussiaDepartment of Microbiology, Faculty of Biology, Lomonosov Moscow State University, Moscow, RussiaKurchatov Complex of Genetic Research, NRC “Kurchatov Institute”, Moscow, RussiaPlant Sciences and the Bioeconomy, Rothamsted Research, Harpenden, United KingdomDepartment of Biosciences, Biotechnologies and Environment, University of Bari, Bari, ItalyDepartment of Biosciences, Biotechnologies and Environment, University of Bari, Bari, ItalyKurchatov Complex of Genetic Research, NRC “Kurchatov Institute”, Moscow, RussiaMendeleev University of Chemical Technology, Moscow, RussiaKurchatov Complex of Genetic Research, NRC “Kurchatov Institute”, Moscow, RussiaDepartment of Biochemistry, Faculty of Biology, Lomonosov Moscow State University, Moscow, RussiaDepartment of Microbiology, Faculty of Biology, Lomonosov Moscow State University, Moscow, RussiaKurchatov Complex of Genetic Research, NRC “Kurchatov Institute”, Moscow, RussiaIn the current study, we report the identification and characterization of the yifK gene product as a novel amino acid carrier in E. coli K-12 cells. Both phenotypic and biochemical analyses showed that YifK acts as a permease specific to L-threonine and, to a lesser extent, L-serine. An assay of the effect of uncouplers and composition of the reaction medium on the transport activity indicates that YifK utilizes a proton motive force to energize substrate uptake. To identify the remaining threonine carriers, we screened a genomic library prepared from the yifK-mutant strain and found that brnQ acts as a multicopy suppressor of the threonine transport defect caused by yifK disruption. Our results indicate that BrnQ is directly involved in threonine uptake as a low-affinity but high-flux transporter, which forms the main entry point when the threonine concentration in the external environment reaches a toxic level. By abolishing YifK and BrnQ activity, we unmasked and quantified the threonine transport activity of the LIV-I branched chain amino acid transport system and demonstrated that LIV-I contributes significantly to total threonine uptake. However, this contribution is likely smaller than that of YifK. We also observed the serine transport activity of LIV-I, which was much lower compared with that of the dedicated SdaC carrier, indicating that LIV-I plays a minor role in the serine uptake. Overall, these findings allow us to propose a comprehensive model of the threonine/serine uptake subsystem in E. coli cells.https://www.frontiersin.org/articles/10.3389/fmicb.2023.1151716/fullEscherichia coliL-serine uptakeL-threonine uptaketransmembrane transportamino acid transporter |
| spellingShingle | Andrey A. Khozov Andrey A. Khozov Dmitrii M. Bubnov Eugeny D. Plisov Tatiana V. Vybornaya Tigran V. Yuzbashev Gennaro Agrimi Eugenia Messina Agnessa A. Stepanova Agnessa A. Stepanova Maxim D. Kudina Natalia V. Alekseeva Alexander I. Netrusov Sergey P. Sineoky A study on L-threonine and L-serine uptake in Escherichia coli K-12 Frontiers in Microbiology Escherichia coli L-serine uptake L-threonine uptake transmembrane transport amino acid transporter |
| title | A study on L-threonine and L-serine uptake in Escherichia coli K-12 |
| title_full | A study on L-threonine and L-serine uptake in Escherichia coli K-12 |
| title_fullStr | A study on L-threonine and L-serine uptake in Escherichia coli K-12 |
| title_full_unstemmed | A study on L-threonine and L-serine uptake in Escherichia coli K-12 |
| title_short | A study on L-threonine and L-serine uptake in Escherichia coli K-12 |
| title_sort | study on l threonine and l serine uptake in escherichia coli k 12 |
| topic | Escherichia coli L-serine uptake L-threonine uptake transmembrane transport amino acid transporter |
| url | https://www.frontiersin.org/articles/10.3389/fmicb.2023.1151716/full |
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