Structural and mechanistic insights into Mycobacterium abscessus as-partate decarboxylase PanD and a pyrazinoic acid-derived inhibitor
Mycobacterium tuberculosis (Mtb) aspartate decarboxylase PanD is required for biosynthesis of the essential cofactor coenzyme A and targeted by the first line drug pyrazinamide (PZA). PZA is a prodrug that is converted by a bacterial amidase into its bioactive form pyrazinoic acid (POA). Employing s...
| Main Authors: | , , , , , , , , , , |
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| Format: | Journal Article |
| Language: | English |
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2022
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| Online Access: | https://hdl.handle.net/10356/163126 |
| _version_ | 1811682500963991552 |
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| author | Saw, Wuan Geok Leow, Chen Yen Harikishore, Amaravadhi Shin, Joon Cole, Malcolm S. Aragaw, Wassihun Wedajo Ragunathan, Priya Hegde, Pooja Aldrich, Courtney C. Dick, Thomas Grüber, Gerhard |
| author2 | School of Biological Sciences |
| author_facet | School of Biological Sciences Saw, Wuan Geok Leow, Chen Yen Harikishore, Amaravadhi Shin, Joon Cole, Malcolm S. Aragaw, Wassihun Wedajo Ragunathan, Priya Hegde, Pooja Aldrich, Courtney C. Dick, Thomas Grüber, Gerhard |
| author_sort | Saw, Wuan Geok |
| collection | NTU |
| description | Mycobacterium tuberculosis (Mtb) aspartate decarboxylase PanD is required for biosynthesis of the essential cofactor coenzyme A and targeted by the first line drug pyrazinamide (PZA). PZA is a prodrug that is converted by a bacterial amidase into its bioactive form pyrazinoic acid (POA). Employing structure-function analyses we previously identified POA-based inhibitors of Mtb PanD showing much improved inhibitory activity against the enzyme. Here, we performed the first structure-function studies on PanD encoded by the non-tuberculous mycobacterial lung pathogen Mycobacterium abscessus (Mab), shedding light on the differences and similarities of Mab and Mtb PanD. Solution X-ray scattering data provided the solution structure of the entire tetrameric Mab PanD, which in comparison to the structure of the derived C-terminal trun-cated Mab PanD1-114 mutant, revealed the orientation of the four flexible C-termini relative to the catalytic core. Enzymatic studies of Mab PanD1-114 explored the essentiality of the C-terminus for catalysis. A library of recombinant Mab PanD mutants based on structural information and PZA/POA resistant PanD mutations in Mtb, illuminated critical residues involved in the substrate tunnel and enzymatic activity. Using our library of POA analogs, we identified (3-(1-naphthamido)pyrazine-2-car-boxylic acid) (analog 2) as the first potent inhibitor of Mab PanD. The inhibitor shows mainly electrostatic- and hydrogen bonding interaction with the target enzyme as explored by isothermal titration calorimetry and confirmed by docking studies. The observed unfavorable entropy indicates that significant conformational changes are involved in the binding process of analog 2 to Mab PanD. In contrast to PZA and POA, which are whole-cell inactive, analog 2 exerts appreciable antibacterial activity against the three subspecies of Mab. |
| first_indexed | 2024-10-01T03:57:50Z |
| format | Journal Article |
| id | ntu-10356/163126 |
| institution | Nanyang Technological University |
| language | English |
| last_indexed | 2024-10-01T03:57:50Z |
| publishDate | 2022 |
| record_format | dspace |
| spelling | ntu-10356/1631262023-02-28T17:12:12Z Structural and mechanistic insights into Mycobacterium abscessus as-partate decarboxylase PanD and a pyrazinoic acid-derived inhibitor Saw, Wuan Geok Leow, Chen Yen Harikishore, Amaravadhi Shin, Joon Cole, Malcolm S. Aragaw, Wassihun Wedajo Ragunathan, Priya Hegde, Pooja Aldrich, Courtney C. Dick, Thomas Grüber, Gerhard School of Biological Sciences Science::Biological sciences::Biochemistry Science::Biological sciences::Biophysics Non-Tuberculous Mycobacteria Mycobacterium Abscessus Pyrazinamide Pyrazinoic Acid Coenzyme A Aspartate Decarboxylase Mycobacterium tuberculosis (Mtb) aspartate decarboxylase PanD is required for biosynthesis of the essential cofactor coenzyme A and targeted by the first line drug pyrazinamide (PZA). PZA is a prodrug that is converted by a bacterial amidase into its bioactive form pyrazinoic acid (POA). Employing structure-function analyses we previously identified POA-based inhibitors of Mtb PanD showing much improved inhibitory activity against the enzyme. Here, we performed the first structure-function studies on PanD encoded by the non-tuberculous mycobacterial lung pathogen Mycobacterium abscessus (Mab), shedding light on the differences and similarities of Mab and Mtb PanD. Solution X-ray scattering data provided the solution structure of the entire tetrameric Mab PanD, which in comparison to the structure of the derived C-terminal trun-cated Mab PanD1-114 mutant, revealed the orientation of the four flexible C-termini relative to the catalytic core. Enzymatic studies of Mab PanD1-114 explored the essentiality of the C-terminus for catalysis. A library of recombinant Mab PanD mutants based on structural information and PZA/POA resistant PanD mutations in Mtb, illuminated critical residues involved in the substrate tunnel and enzymatic activity. Using our library of POA analogs, we identified (3-(1-naphthamido)pyrazine-2-car-boxylic acid) (analog 2) as the first potent inhibitor of Mab PanD. The inhibitor shows mainly electrostatic- and hydrogen bonding interaction with the target enzyme as explored by isothermal titration calorimetry and confirmed by docking studies. The observed unfavorable entropy indicates that significant conformational changes are involved in the binding process of analog 2 to Mab PanD. In contrast to PZA and POA, which are whole-cell inactive, analog 2 exerts appreciable antibacterial activity against the three subspecies of Mab. Ministry of Education (MOE) National Research Foundation (NRF) Submitted/Accepted version Research reported in this publication is supported by the National Institute of Allergy and Infectious Diseases of the National Institutes of Health under Award Numbers 2R01AI106398-05 (T.D., (T.D., (T.D., C.A.,C.A., G.G.) G.G.) . The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. The studies are also supported by the National Research Founda-tion (NRF) Singapore, NRF Competitive Research Programme (CRP), Grant Award Number NRF–CRP18–2017–01 (G.G., T.D.), and the Singapore Ministry of Education (MOE) Academic Research Fund Tier 1 (RG107/20) to G.G. 2022-11-24T05:39:13Z 2022-11-24T05:39:13Z 2022 Journal Article Saw, W. G., Leow, C. Y., Harikishore, A., Shin, J., Cole, M. S., Aragaw, W. W., Ragunathan, P., Hegde, P., Aldrich, C. C., Dick, T. & Grüber, G. (2022). Structural and mechanistic insights into Mycobacterium abscessus as-partate decarboxylase PanD and a pyrazinoic acid-derived inhibitor. ACS Infectious Disease, 8(7), 1324-1335. https://dx.doi.org/10.1021/acsinfecdis.2c00133 2373-8227 https://hdl.handle.net/10356/163126 10.1021/acsinfecdis.2c00133 7 8 1324 1335 en NRF-CRP18-2017- 01 RG107/20 ACS Infectious Disease This document is the Accepted Manuscript version of a Published Work that appeared in final form in ACS Infectious Diseases, copyright © 2022 American Chemical Society, after peer review and technical editing by the publisher. To access the final edited and published work see https://doi.org/10.1021/acsinfecdis.2c00133. application/pdf |
| spellingShingle | Science::Biological sciences::Biochemistry Science::Biological sciences::Biophysics Non-Tuberculous Mycobacteria Mycobacterium Abscessus Pyrazinamide Pyrazinoic Acid Coenzyme A Aspartate Decarboxylase Saw, Wuan Geok Leow, Chen Yen Harikishore, Amaravadhi Shin, Joon Cole, Malcolm S. Aragaw, Wassihun Wedajo Ragunathan, Priya Hegde, Pooja Aldrich, Courtney C. Dick, Thomas Grüber, Gerhard Structural and mechanistic insights into Mycobacterium abscessus as-partate decarboxylase PanD and a pyrazinoic acid-derived inhibitor |
| title | Structural and mechanistic insights into Mycobacterium abscessus as-partate decarboxylase PanD and a pyrazinoic acid-derived inhibitor |
| title_full | Structural and mechanistic insights into Mycobacterium abscessus as-partate decarboxylase PanD and a pyrazinoic acid-derived inhibitor |
| title_fullStr | Structural and mechanistic insights into Mycobacterium abscessus as-partate decarboxylase PanD and a pyrazinoic acid-derived inhibitor |
| title_full_unstemmed | Structural and mechanistic insights into Mycobacterium abscessus as-partate decarboxylase PanD and a pyrazinoic acid-derived inhibitor |
| title_short | Structural and mechanistic insights into Mycobacterium abscessus as-partate decarboxylase PanD and a pyrazinoic acid-derived inhibitor |
| title_sort | structural and mechanistic insights into mycobacterium abscessus as partate decarboxylase pand and a pyrazinoic acid derived inhibitor |
| topic | Science::Biological sciences::Biochemistry Science::Biological sciences::Biophysics Non-Tuberculous Mycobacteria Mycobacterium Abscessus Pyrazinamide Pyrazinoic Acid Coenzyme A Aspartate Decarboxylase |
| url | https://hdl.handle.net/10356/163126 |
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