Mycobacterial F420H2-Dependent Reductases Promiscuously Reduce Diverse Compounds through a Common Mechanism
An unusual aspect of actinobacterial metabolism is the use of the redox cofactor F420. Studies have shown that actinobacterial F420H2-dependent reductases promiscuously hydrogenate diverse organic compounds in biodegradative and biosynthetic processes. These enzymes therefore represent promising can...
| Main Authors: | , , , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2017-05-01
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| Series: | Frontiers in Microbiology |
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| Online Access: | http://journal.frontiersin.org/article/10.3389/fmicb.2017.01000/full |
| _version_ | 1818366261905588224 |
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| author | Chris Greening Chris Greening Thanavit Jirapanjawat Thanavit Jirapanjawat Thanavit Jirapanjawat Shahana Afroze Shahana Afroze Blair Ney Blair Ney Blair Ney Colin Scott Gunjan Pandey Brendon M. Lee Brendon M. Lee Robyn J. Russell Colin J. Jackson John G. Oakeshott Matthew C. Taylor Andrew C. Warden |
| author_facet | Chris Greening Chris Greening Thanavit Jirapanjawat Thanavit Jirapanjawat Thanavit Jirapanjawat Shahana Afroze Shahana Afroze Blair Ney Blair Ney Blair Ney Colin Scott Gunjan Pandey Brendon M. Lee Brendon M. Lee Robyn J. Russell Colin J. Jackson John G. Oakeshott Matthew C. Taylor Andrew C. Warden |
| author_sort | Chris Greening |
| collection | DOAJ |
| description | An unusual aspect of actinobacterial metabolism is the use of the redox cofactor F420. Studies have shown that actinobacterial F420H2-dependent reductases promiscuously hydrogenate diverse organic compounds in biodegradative and biosynthetic processes. These enzymes therefore represent promising candidates for next-generation industrial biocatalysts. In this work, we undertook the first broad survey of these enzymes as potential industrial biocatalysts by exploring the extent, as well as mechanistic and structural bases, of their substrate promiscuity. We expressed and purified 11 enzymes from seven subgroups of the flavin/deazaflavin oxidoreductase (FDOR) superfamily (A1, A2, A3, B1, B2, B3, B4) from the model soil actinobacterium Mycobacterium smegmatis. These enzymes reduced compounds from six chemical classes, including fundamental monocycles such as a cyclohexenone, a dihydropyran, and pyrones, as well as more complex quinone, coumarin, and arylmethane compounds. Substrate range and reduction rates varied between the enzymes, with the A1, A3, and B1 groups exhibiting greatest promiscuity. Molecular docking studies suggested that structurally diverse compounds are accommodated in the large substrate-binding pocket of the most promiscuous FDOR through hydrophobic interactions with conserved aromatic residues and the isoalloxazine headgroup of F420H2. Liquid chromatography-mass spectrometry (LC/MS) and gas chromatography-mass spectrometry (GC/MS) analysis of derivatized reaction products showed reduction occurred through a common mechanism involving hydride transfer from F420H- to the electron-deficient alkene groups of substrates. Reduction occurs when the hydride donor (C5 of F420H-) is proximal to the acceptor (electrophilic alkene of the substrate). These findings suggest that engineered actinobacterial F420H2-dependent reductases are promising novel biocatalysts for the facile transformation of a wide range of α,β-unsaturated compounds. |
| first_indexed | 2024-12-13T22:33:21Z |
| format | Article |
| id | doaj.art-9c3eee481f1c46efab8c6eb800289757 |
| institution | Directory Open Access Journal |
| issn | 1664-302X |
| language | English |
| last_indexed | 2024-12-13T22:33:21Z |
| publishDate | 2017-05-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Microbiology |
| spelling | doaj.art-9c3eee481f1c46efab8c6eb8002897572022-12-21T23:29:02ZengFrontiers Media S.A.Frontiers in Microbiology1664-302X2017-05-01810.3389/fmicb.2017.01000270757Mycobacterial F420H2-Dependent Reductases Promiscuously Reduce Diverse Compounds through a Common MechanismChris Greening0Chris Greening1Thanavit Jirapanjawat2Thanavit Jirapanjawat3Thanavit Jirapanjawat4Shahana Afroze5Shahana Afroze6Blair Ney7Blair Ney8Blair Ney9Colin Scott10Gunjan Pandey11Brendon M. Lee12Brendon M. Lee13Robyn J. Russell14Colin J. Jackson15John G. Oakeshott16Matthew C. Taylor17Andrew C. Warden18Land and Water Flagship, The Commonwealth Scientific and Industrial Research Organisation, ActonACT, AustraliaSchool of Biological Sciences, Monash University, ClaytonVIC, AustraliaLand and Water Flagship, The Commonwealth Scientific and Industrial Research Organisation, ActonACT, AustraliaSchool of Biological Sciences, Monash University, ClaytonVIC, AustraliaResearch School of Chemistry, Australian National University, ActonACT, AustraliaLand and Water Flagship, The Commonwealth Scientific and Industrial Research Organisation, ActonACT, AustraliaResearch School of Chemistry, Australian National University, ActonACT, AustraliaLand and Water Flagship, The Commonwealth Scientific and Industrial Research Organisation, ActonACT, AustraliaSchool of Biological Sciences, Monash University, ClaytonVIC, AustraliaResearch School of Chemistry, Australian National University, ActonACT, AustraliaLand and Water Flagship, The Commonwealth Scientific and Industrial Research Organisation, ActonACT, AustraliaLand and Water Flagship, The Commonwealth Scientific and Industrial Research Organisation, ActonACT, AustraliaLand and Water Flagship, The Commonwealth Scientific and Industrial Research Organisation, ActonACT, AustraliaResearch School of Chemistry, Australian National University, ActonACT, AustraliaLand and Water Flagship, The Commonwealth Scientific and Industrial Research Organisation, ActonACT, AustraliaResearch School of Chemistry, Australian National University, ActonACT, AustraliaLand and Water Flagship, The Commonwealth Scientific and Industrial Research Organisation, ActonACT, AustraliaLand and Water Flagship, The Commonwealth Scientific and Industrial Research Organisation, ActonACT, AustraliaLand and Water Flagship, The Commonwealth Scientific and Industrial Research Organisation, ActonACT, AustraliaAn unusual aspect of actinobacterial metabolism is the use of the redox cofactor F420. Studies have shown that actinobacterial F420H2-dependent reductases promiscuously hydrogenate diverse organic compounds in biodegradative and biosynthetic processes. These enzymes therefore represent promising candidates for next-generation industrial biocatalysts. In this work, we undertook the first broad survey of these enzymes as potential industrial biocatalysts by exploring the extent, as well as mechanistic and structural bases, of their substrate promiscuity. We expressed and purified 11 enzymes from seven subgroups of the flavin/deazaflavin oxidoreductase (FDOR) superfamily (A1, A2, A3, B1, B2, B3, B4) from the model soil actinobacterium Mycobacterium smegmatis. These enzymes reduced compounds from six chemical classes, including fundamental monocycles such as a cyclohexenone, a dihydropyran, and pyrones, as well as more complex quinone, coumarin, and arylmethane compounds. Substrate range and reduction rates varied between the enzymes, with the A1, A3, and B1 groups exhibiting greatest promiscuity. Molecular docking studies suggested that structurally diverse compounds are accommodated in the large substrate-binding pocket of the most promiscuous FDOR through hydrophobic interactions with conserved aromatic residues and the isoalloxazine headgroup of F420H2. Liquid chromatography-mass spectrometry (LC/MS) and gas chromatography-mass spectrometry (GC/MS) analysis of derivatized reaction products showed reduction occurred through a common mechanism involving hydride transfer from F420H- to the electron-deficient alkene groups of substrates. Reduction occurs when the hydride donor (C5 of F420H-) is proximal to the acceptor (electrophilic alkene of the substrate). These findings suggest that engineered actinobacterial F420H2-dependent reductases are promising novel biocatalysts for the facile transformation of a wide range of α,β-unsaturated compounds.http://journal.frontiersin.org/article/10.3389/fmicb.2017.01000/fullF420redoxbiocatalysispromiscuitybiodegradationMycobacterium |
| spellingShingle | Chris Greening Chris Greening Thanavit Jirapanjawat Thanavit Jirapanjawat Thanavit Jirapanjawat Shahana Afroze Shahana Afroze Blair Ney Blair Ney Blair Ney Colin Scott Gunjan Pandey Brendon M. Lee Brendon M. Lee Robyn J. Russell Colin J. Jackson John G. Oakeshott Matthew C. Taylor Andrew C. Warden Mycobacterial F420H2-Dependent Reductases Promiscuously Reduce Diverse Compounds through a Common Mechanism Frontiers in Microbiology F420 redox biocatalysis promiscuity biodegradation Mycobacterium |
| title | Mycobacterial F420H2-Dependent Reductases Promiscuously Reduce Diverse Compounds through a Common Mechanism |
| title_full | Mycobacterial F420H2-Dependent Reductases Promiscuously Reduce Diverse Compounds through a Common Mechanism |
| title_fullStr | Mycobacterial F420H2-Dependent Reductases Promiscuously Reduce Diverse Compounds through a Common Mechanism |
| title_full_unstemmed | Mycobacterial F420H2-Dependent Reductases Promiscuously Reduce Diverse Compounds through a Common Mechanism |
| title_short | Mycobacterial F420H2-Dependent Reductases Promiscuously Reduce Diverse Compounds through a Common Mechanism |
| title_sort | mycobacterial f420h2 dependent reductases promiscuously reduce diverse compounds through a common mechanism |
| topic | F420 redox biocatalysis promiscuity biodegradation Mycobacterium |
| url | http://journal.frontiersin.org/article/10.3389/fmicb.2017.01000/full |
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