دورية أكاديمية
The NADH recycling enzymes TsaC and TsaD regenerate reducing equivalents for Rieske oxygenase chemistry.
| العنوان: | The NADH recycling enzymes TsaC and TsaD regenerate reducing equivalents for Rieske oxygenase chemistry. |
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| المؤلفون: | Tian J; Department of Chemistry, University of Michigan, Ann Arbor, Michigan, USA., Boggs DG; Department of Chemistry, University of Michigan, Ann Arbor, Michigan, USA., Donnan PH; Department of Chemistry, University of Michigan, Ann Arbor, Michigan, USA., Barroso GT; Department of Chemistry, University of Michigan, Ann Arbor, Michigan, USA., Garcia AA; Department of Chemistry, University of Michigan, Ann Arbor, Michigan, USA., Dowling DP; Department of Chemistry, University of Massachusetts Boston, Boston, Massachusetts, USA., Buss JA; Department of Chemistry, University of Michigan, Ann Arbor, Michigan, USA., Bridwell-Rabb J; Department of Chemistry, University of Michigan, Ann Arbor, Michigan, USA. Electronic address: jebridwe@umich.edu. |
| المصدر: | The Journal of biological chemistry [J Biol Chem] 2023 Oct; Vol. 299 (10), pp. 105222. Date of Electronic Publication: 2023 Sep 09. |
| نوع المنشور: | Journal Article; Research Support, N.I.H., Extramural |
| اللغة: | English |
| بيانات الدورية: | Publisher: Elsevier Inc. on behalf of American Society for Biochemistry and Molecular Biology Country of Publication: United States NLM ID: 2985121R Publication Model: Print-Electronic Cited Medium: Internet ISSN: 1083-351X (Electronic) Linking ISSN: 00219258 NLM ISO Abbreviation: J Biol Chem Subsets: MEDLINE |
| أسماء مطبوعة: | Publication: 2021- : [New York, NY] : Elsevier Inc. on behalf of American Society for Biochemistry and Molecular Biology Original Publication: Baltimore, MD : American Society for Biochemistry and Molecular Biology |
| مواضيع طبية MeSH: | Oxygenases*/metabolism , Comamonas testosteroni*/enzymology , Comamonas testosteroni*/genetics , Bacterial Proteins*/chemistry , Bacterial Proteins*/genetics , Bacterial Proteins*/metabolism, Aldehyde Dehydrogenase/metabolism ; NAD/metabolism ; Substrate Specificity ; Nonheme Iron Proteins/chemistry ; Nonheme Iron Proteins/genetics ; Nonheme Iron Proteins/metabolism ; Recombinant Proteins/chemistry ; Recombinant Proteins/genetics ; Recombinant Proteins/metabolism ; Protein Structure, Tertiary ; Models, Molecular ; Protein Stability ; Computational Biology |
| مستخلص: | Many microorganisms use both biological and nonbiological molecules as sources of carbon and energy. This resourcefulness means that some microorganisms have mechanisms to assimilate pollutants found in the environment. One such organism is Comamonas testosteroni, which metabolizes 4-methylbenzenesulfonate and 4-methylbenzoate using the TsaMBCD pathway. TsaM is a Rieske oxygenase, which in concert with the reductase TsaB consumes a molar equivalent of NADH. Following this step, the annotated short-chain dehydrogenase/reductase and aldehyde dehydrogenase enzymes TsaC and TsaD each regenerate a molar equivalent of NADH. This co-occurrence ameliorates the need for stoichiometric addition of reducing equivalents and thus represents an attractive strategy for integration of Rieske oxygenase chemistry into biocatalytic applications. Therefore, in this work, to overcome the lack of information regarding NADH recycling enzymes that function in partnership with Rieske non-heme iron oxygenases (Rieske oxygenases), we solved the X-ray crystal structure of TsaC to a resolution of 2.18 Å. Using this structure, a series of substrate analog and protein variant combination reactions, and differential scanning fluorimetry experiments, we identified active site features involved in binding NAD + and controlling substrate specificity. Further in vitro enzyme cascade experiments demonstrated the efficient TsaC- and TsaD-mediated regeneration of NADH to support Rieske oxygenase chemistry. Finally, through in-depth bioinformatic analyses, we illustrate the widespread co-occurrence of Rieske oxygenases with TsaC-like enzymes. This work thus demonstrates the utility of these NADH recycling enzymes and identifies a library of short-chain dehydrogenase/reductase enzyme prospects that can be used in Rieske oxygenase pathways for in situ regeneration of NADH. Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article. (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.) |
| معلومات مُعتمدة: | R35 GM138271 United States GM NIGMS NIH HHS |
| فهرسة مساهمة: | Keywords: NAD; Rieske oxygenase; X-ray crystallography; catabolism; enzyme catalysis; enzyme degradation; enzyme kinetics; pollutants; short-chain dehydrogenase reductase |
| المشرفين على المادة: | EC 1.2.1.3 (Aldehyde Dehydrogenase) 0U46U6E8UK (NAD) EC 1.13.- (Oxygenases) 0 (Bacterial Proteins) 0 (Nonheme Iron Proteins) 0 (Recombinant Proteins) |
| تواريخ الأحداث: | Date Created: 20230906 Date Completed: 20240214 Latest Revision: 20240624 |
| رمز التحديث: | 20240624 |
| مُعرف محوري في PubMed: | PMC10579966 |
| DOI: | 10.1016/j.jbc.2023.105222 |
| PMID: | 37673337 |
| قاعدة البيانات: | MEDLINE |
Full Text Finder | تدمد: | 1083-351X |
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| DOI: | 10.1016/j.jbc.2023.105222 |