Re-investigation of in vitro activity of acetohydroxyacid synthase I holoenzyme from Escherichia coli
Copyright © 2024. Published by Elsevier Inc..
Acetohydroxyacid synthase (AHAS) is one of the key enzymes of the biosynthesis of branched-chain amino acids, it is also an effective target for the screening of herbicides and antibiotics. In this study we present a method for preparing Escherichia coli AHAS I holoenzyme (EcAHAS I) with exceptional stability, which provides a solid ground for us to re-investigate the in vitro catalytic properties of the protein. The results show EcAHAS I synthesized in this way exhibits similar function to Bacillus subtilis acetolactate synthase in its catalysis with pyruvate and 2-ketobutyrate (2-KB) as dual-substrate, producing four 2-hydroxy-3-ketoacids including (S)-2-acetolactate, (S)-2-aceto-2-hydroxybutyrate, (S)-2-propionyllactate, and (S)-2-propionyl-2-hydroxybutyrate. Quantification of the reaction indicates that the two substrates almost totally consume, and compound (S)-2-aceto-2- hydroxybutyrate forms in the highest yield among the four major products. Moreover, the protein also condenses two molecules of 2-KB to furnish (S)-2-propionyl-2-hydroxybutyrate. Further exploration manifests that EcAHAS I ligates pyruvate/2-KB and nitrosobenzene to generate two arylhydroxamic acids N-hydroxy-N-phenylacetamide and N-hydroxy-N-phenyl- propionamide. These findings enhance our comprehension of the catalytic characteristics of EcAHAS I. Furthermore, the application of this enzyme as a catalyst in construction of C-N bonds displays promising potential.
| Medienart: |
E-Artikel |
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| Erscheinungsjahr: |
2024 |
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| Erschienen: |
2024 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:754 |
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| Enthalten in: |
Archives of biochemistry and biophysics - 754(2024) vom: 29. Apr., Seite 109962 |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Wang, Hai-Ling [VerfasserIn] |
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| Links: |
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| Anmerkungen: |
Date Completed 01.04.2024 Date Revised 01.04.2024 published: Print-Electronic Citation Status MEDLINE |
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| doi: |
10.1016/j.abb.2024.109962 |
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| Förderinstitution / Projekttitel: |
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| 520 | |a Acetohydroxyacid synthase (AHAS) is one of the key enzymes of the biosynthesis of branched-chain amino acids, it is also an effective target for the screening of herbicides and antibiotics. In this study we present a method for preparing Escherichia coli AHAS I holoenzyme (EcAHAS I) with exceptional stability, which provides a solid ground for us to re-investigate the in vitro catalytic properties of the protein. The results show EcAHAS I synthesized in this way exhibits similar function to Bacillus subtilis acetolactate synthase in its catalysis with pyruvate and 2-ketobutyrate (2-KB) as dual-substrate, producing four 2-hydroxy-3-ketoacids including (S)-2-acetolactate, (S)-2-aceto-2-hydroxybutyrate, (S)-2-propionyllactate, and (S)-2-propionyl-2-hydroxybutyrate. Quantification of the reaction indicates that the two substrates almost totally consume, and compound (S)-2-aceto-2- hydroxybutyrate forms in the highest yield among the four major products. Moreover, the protein also condenses two molecules of 2-KB to furnish (S)-2-propionyl-2-hydroxybutyrate. Further exploration manifests that EcAHAS I ligates pyruvate/2-KB and nitrosobenzene to generate two arylhydroxamic acids N-hydroxy-N-phenylacetamide and N-hydroxy-N-phenyl- propionamide. These findings enhance our comprehension of the catalytic characteristics of EcAHAS I. Furthermore, the application of this enzyme as a catalyst in construction of C-N bonds displays promising potential | ||
| 650 | 4 | |a Journal Article | |
| 650 | 4 | |a Acetohydroxyacid synthase | |
| 650 | 4 | |a Enzyme catalysis | |
| 650 | 4 | |a Enzyme kinetics | |
| 650 | 4 | |a Escherichia coli | |
| 650 | 4 | |a Preparation | |
| 650 | 7 | |a Acetolactate Synthase |2 NLM | |
| 650 | 7 | |a EC 2.2.1.6 |2 NLM | |
| 650 | 7 | |a Glycogen Synthase |2 NLM | |
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| 650 | 7 | |a alpha-aceto-alpha-hydroxybutyrate |2 NLM | |
| 650 | 7 | |a 3142-65-2 |2 NLM | |
| 650 | 7 | |a Hydroxybutyrates |2 NLM | |
| 650 | 7 | |a Pyruvates |2 NLM | |
| 650 | 7 | |a Holoenzymes |2 NLM | |
| 700 | 1 | |a Sun, Hui-Peng |e verfasserin |4 aut | |
| 700 | 1 | |a Zheng, Pei-Rong |e verfasserin |4 aut | |
| 700 | 1 | |a Cheng, Rui-Tong |e verfasserin |4 aut | |
| 700 | 1 | |a Liu, Zhi-Wen |e verfasserin |4 aut | |
| 700 | 1 | |a Yuan, Heng |e verfasserin |4 aut | |
| 700 | 1 | |a Gao, Wen-Yun |e verfasserin |4 aut | |
| 700 | 1 | |a Li, Heng |e verfasserin |4 aut | |
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