Redesign of ultrasensitive and robust RecA gene circuit to sense DNA damage

Summary SOS box of the recA promoter, PVRecA from Vibrio natriegens was characterized, cloned and expressed in a probiotic strain E. coli Nissle 1917. This promoter was then rationally engineered according to predicted interactions between LexA repressor and PVRecA. The redesigned PVRecA‐AT promoter...

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Autores principales:	Jack X. Chen, Boon Lim, Harrison Steel, Yizhi Song, Mengmeng Ji, Wei E. Huang
Formato:	article
Lenguaje:	EN
Publicado:	Wiley 2021
Materias:	Biotechnology TP248.13-248.65
Acceso en línea:	https://doaj.org/article/1d917eaa409b4fd6825382f017ee98bb
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spelling	oai:doaj.org-article:1d917eaa409b4fd6825382f017ee98bb2021-11-18T15:39:52ZRedesign of ultrasensitive and robust RecA gene circuit to sense DNA damage1751-791510.1111/1751-7915.13767https://doaj.org/article/1d917eaa409b4fd6825382f017ee98bb2021-11-01T00:00:00Zhttps://doi.org/10.1111/1751-7915.13767https://doaj.org/toc/1751-7915Summary SOS box of the recA promoter, PVRecA from Vibrio natriegens was characterized, cloned and expressed in a probiotic strain E. coli Nissle 1917. This promoter was then rationally engineered according to predicted interactions between LexA repressor and PVRecA. The redesigned PVRecA‐AT promoter showed a sensitive and robust response to DNA damage induced by UV and genotoxic compounds. Rational design of PVRecA coupled to an amplification gene circuit increased circuit output amplitude 4.3‐fold in response to a DNA damaging compound mitomycin C. A TetR‐based negative feedback loop was added to the PVRecA‐AT amplifier to achieve a robust SOS system, resistant to environmental fluctuations in parameters including pH, temperature, oxygen and nutrient conditions. We found that E. coli Nissle 1917 with optimized PVRecA‐AT adapted to UV exposure and increased SOS response 128‐fold over 40 h cultivation in turbidostat mini‐reactor. We also showed the potential of this PVRecA‐AT system as an optogenetic actuator, which can be controlled spatially through UV radiation. We demonstrated that the optimized SOS responding gene circuits were able to detect carcinogenic biomarker molecules with clinically relevant concentrations. The ultrasensitive SOS gene circuits in probiotic E. coli Nissle 1917 would be potentially useful for bacterial diagnosis.Jack X. ChenBoon LimHarrison SteelYizhi SongMengmeng JiWei E. HuangWileyarticleBiotechnologyTP248.13-248.65ENMicrobial Biotechnology, Vol 14, Iss 6, Pp 2481-2496 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Biotechnology TP248.13-248.65
spellingShingle	Biotechnology TP248.13-248.65 Jack X. Chen Boon Lim Harrison Steel Yizhi Song Mengmeng Ji Wei E. Huang Redesign of ultrasensitive and robust RecA gene circuit to sense DNA damage
description	Summary SOS box of the recA promoter, PVRecA from Vibrio natriegens was characterized, cloned and expressed in a probiotic strain E. coli Nissle 1917. This promoter was then rationally engineered according to predicted interactions between LexA repressor and PVRecA. The redesigned PVRecA‐AT promoter showed a sensitive and robust response to DNA damage induced by UV and genotoxic compounds. Rational design of PVRecA coupled to an amplification gene circuit increased circuit output amplitude 4.3‐fold in response to a DNA damaging compound mitomycin C. A TetR‐based negative feedback loop was added to the PVRecA‐AT amplifier to achieve a robust SOS system, resistant to environmental fluctuations in parameters including pH, temperature, oxygen and nutrient conditions. We found that E. coli Nissle 1917 with optimized PVRecA‐AT adapted to UV exposure and increased SOS response 128‐fold over 40 h cultivation in turbidostat mini‐reactor. We also showed the potential of this PVRecA‐AT system as an optogenetic actuator, which can be controlled spatially through UV radiation. We demonstrated that the optimized SOS responding gene circuits were able to detect carcinogenic biomarker molecules with clinically relevant concentrations. The ultrasensitive SOS gene circuits in probiotic E. coli Nissle 1917 would be potentially useful for bacterial diagnosis.
format	article
author	Jack X. Chen Boon Lim Harrison Steel Yizhi Song Mengmeng Ji Wei E. Huang
author_facet	Jack X. Chen Boon Lim Harrison Steel Yizhi Song Mengmeng Ji Wei E. Huang
author_sort	Jack X. Chen
title	Redesign of ultrasensitive and robust RecA gene circuit to sense DNA damage
title_short	Redesign of ultrasensitive and robust RecA gene circuit to sense DNA damage
title_full	Redesign of ultrasensitive and robust RecA gene circuit to sense DNA damage
title_fullStr	Redesign of ultrasensitive and robust RecA gene circuit to sense DNA damage
title_full_unstemmed	Redesign of ultrasensitive and robust RecA gene circuit to sense DNA damage
title_sort	redesign of ultrasensitive and robust reca gene circuit to sense dna damage
publisher	Wiley
publishDate	2021
url	https://doaj.org/article/1d917eaa409b4fd6825382f017ee98bb
work_keys_str_mv	AT jackxchen redesignofultrasensitiveandrobustrecagenecircuittosensednadamage AT boonlim redesignofultrasensitiveandrobustrecagenecircuittosensednadamage AT harrisonsteel redesignofultrasensitiveandrobustrecagenecircuittosensednadamage AT yizhisong redesignofultrasensitiveandrobustrecagenecircuittosensednadamage AT mengmengji redesignofultrasensitiveandrobustrecagenecircuittosensednadamage AT weiehuang redesignofultrasensitiveandrobustrecagenecircuittosensednadamage
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Redesign of ultrasensitive and robust RecA gene circuit to sense DNA damage

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