Genetic biosensors for small-molecule products: Design and applications in high-throughput screening

doi:10.1007/s11705-017-1629-z

Front. Chem. Sci. Eng.

2017, Vol. 11

Issue (1) : 15-26 https://doi.org/10.1007/s11705-017-1629-z

REVIEW ARTICLE

Genetic biosensors for small-molecule products: Design and applications in high-throughput screening

Qingzhuo Wang^1,²,Shuang-Yan Tang³(

),Sheng Yang^1,⁴(

)

¹. Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Shanghai 200025, China
². University of the Chinese Academy of Sciences, Beijing 100049, China
³. CAS Key Laboratory of Microbial Physiological and Metebolic Engineering, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China
⁴. Jiangsu National Synergetic Innovation Center for Advanced Materials, Nanjing Tech University, Nanjing 211816, China

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Abstract

Overproduction of small-molecule chemicals using engineered microbial cells has greatly reduced the production cost and promoted environmental protection. Notably, the rapid and sensitive evaluation of the in vivo concentrations of the desired products greatly facilitates the optimization process of cell factories. For this purpose, many genetic components have been adapted into in vivo biosensors of small molecules, which couple the intracellular concentrations of small molecules to easily detectable readouts such as fluorescence, absorbance, and cell growth. Such biosensors allow a high-throughput screening of the small-molecule products, and can be roughly classified as protein-based and RNA-based biosensors. This review summarizes the recent developments in the design and applications of biosensors for small-molecule products.

Keywords biosensor small molecule product transcription factor riboswitch high-throughput screening

PACS:

Fund:

Corresponding Author(s): Shuang-Yan Tang,Sheng Yang

Just Accepted Date: 24 January 2017 Online First Date: 28 February 2017 Issue Date: 17 March 2017

Cite this article:

Qingzhuo Wang,Shuang-Yan Tang,Sheng Yang. Genetic biosensors for small-molecule products: Design and applications in high-throughput screening[J]. Front. Chem. Sci. Eng., 2017, 11(1): 15-26.

URL:

https://academic.hep.com.cn/fcse/EN/10.1007/s11705-017-1629-z
https://academic.hep.com.cn/fcse/EN/Y2017/V11/I1/15

Fig.1 Ladder of library screening and selection.

The choice of screening method dramatically impacting the throughput; genetic biosensors convert the in vivo concentrations of small-molecule products into easily detectable readouts and improve the throughput of hyper-producers screening

Fig.2 Types of genetically-encoded metabolite biosensors.

The schematic representation of (a, b, c) protein-based biosensors, and (d, e) RNA-based biosensors, and their respective modes of action. The OFF state and small metabolite-triggered ON state of the biosensors are depicted. (a) Transcription factor-based biosensors; (b) yeast three-hybrid biosensors; (c) FRET biosensors; (d) a RNA biosensor based on transcription modulation; (c) hammerhead ribozyme-based riboswitches

Type of target molecule	Target molecule	Type of biosensor	Host cells	Screen method	Reporter	Sensor module (source)	Effect module (source)	Ref.
Amino acids	$L$ -Lysine	RNA	Escherichia coli	Growth selection	TetA-GFP	5'-UTR region of lysC gene (Escherichia coli)		[42,67]
	$L$ -Tryptophan	RNA	Escherichia coli	Growth selection	TetA-GFP	5'UTR region of lysC gene (Escherichia coli)		[42]
	$L$ -Lysine	TF	Corynebacterium glutamicum	FACS	EYFP	LysG (Corynebacterium glutamicum)	Promoter P_lysE	[28,58]
	$L$ -Arginine	TF	Escherichia coli	?	EYFP	ArgP (Escherichia coli)	Promoter P_argO	[28]
	$L$ -Serine	TF	Corynebacterium glutamicum	?	EYFP	NCgl0581 (Escherichia coli)	Promoter P_NCgl0580	[28]
	$L$ -Arginine, $L$ -Histidine	TF	Corynebacterium glutamicum	FACS	EYFP	LysG (Corynebacterium glutamicum)	Promoter P_lysE	[28,59]
	$L$ -Methionine, $L$ -Leucine, $L$ -Isoleucine	TF	Corynebacterium glutamicum	FACS	EYFP	Lrp (Corynebacterium glutamicum)	Promoter P_brnF	[61]
	$L$ -Valine	TF	Corynebacterium glutamicum	FACS	EYFP	Lrp (Corynebacterium glutamicum)	Promoter P_brnF	[61,62]
	$L$ -Tyrosine	TF	Escherichia coli	?	MutD5-mCherry	TyrR (Escherichia coli)	Promoter P_aroF	[86]
	$L$ -Leucine	FRET	Escherichia coli	?	CFP+YFP	LivK (Escherichia coli)		[66]
	$L$ -Methionine	FRET	Escherichia coli/Saccharomyces cerevisiae	?	CFP+YFP	MetN (Escherichia coli)		[65]
	$L$ -Threonine	Stress-response promoter	Escherichia coli	FACS	EGFP	Fusion promoter cysJHp (Escherichia coli)		[68]
	$L$ -Phenylalanine	Stress-response promoter	Escherichia coli	FACS	GFP-Mut2	Promoter P_mtr(Escherichia coli)		[70]
	$L$ -Lysine	Stress-response promoter	Escherichia coli	FACS	EGFP	Promoter pA (Corynebacterium glutamicum)		[87] [88]
Organic acids	Benzoate, 2-Hydroxybenzoate	TF	Escherichia coli	Growth selection	LacZ/TetA	NahR (Pseudomonas putida)	Promoter P_sal	[74]
	Benzoate	TF	Escherichia coli	96-well plates	GFP	BenR (Metagenome)	Promoter P_BenA	[73]
	Mevalonate	TF	Escherichia coli	FACS/visual observation	GFPuv/LacZ	AraC-mev	Promoter P_BAD	[78]
	Triacetic acid lactone	TF	Escherichia coli	FACS /visual observation	GFPuv/LacZ	AraC-TAL	Promoter P_BAD	[79]
	Ectoine	TF	Escherichia coli	FACS	GFPuv	AraC-ect	Promoter P_BAD	[76]
	Adipate	TF	Escherichia coli	?	TetA-GFP	PcaR (Pseudomonas putida)	Promoter P_caIJ	[71]
	Glucaric acid	TF	Escherichia coli	Growth selection	TolC	CdaR (Escherichia coli)	Promoter gudPp	[75]
	Succinate	Two-component system	Escherichia coli	?	TetA	DcuR/DcuS (Escherichia coli)	Promoter P_dctA	[71]
	Medium-chain fatty acids (C8-C12)	G-protein coupled receptor sensor	Saccharomyces cerevisiae	?	GFP	Olfactory receptor OR1G1/ Free fatty acid receptor GPR40	TF-promoter pairs	[77]
Alcohols	Linear alcohols ( $L$ -butanol, $L$ -pentanol, $L$ -hexanol)	TF	Escherichia coli	Growth selection	TetA-GFP	BomR (Thauera butanivorans)	Promoter P_BMO	[71]
	Branched-chain alcohols (2-Methyl- $L$ -propanol, 2-methyl- $L$ -butanol, 3-methyl- $L$ -butanol)	TF	Escherichia coli	?	TetA-GFP	BomR (Thauera butanivorans)	Promoter P_BMO	[71]
	1,4-Butanediol	TF	Escherichia coli	?	TetA-GFP	BomR (Thauera butanivorans)	Promoter P_BMO	[71]
	1,4-Butanediol	Stress-response promoter	Escherichia coli	?	GFP	Promoter P_yhjX		[72]
Flavonoids	Kaempferol	TF	Escherichia coli	96-Well plates	GFP	QdoR (Bacillus subtilis)	Promoter P_qdoI	[81]
	Quercetin	TF	Escherichia coli	?	GFP	QdoR (Bacillus subtilis)	Promoter P_qdoI	[81]
	Naringenin	TF	Escherichia coli	?	GFP	FdeR (Herbaspirillum seropedicae)	Promoter P_fdeA	[81]
	Naringenin	TF	Escherichia coli	Growth selection	TolC	TtgR (Pseudomonas putida)	Promoter ttgAp	[75]
Theophylline	Theophylline	RNA	Escherichia coli	Growth selection	Thymidylate synthase	Synthetic aptamer mTCT8-4 aptamer	Group I aptazyme (Bacteriophage T4)	[85]
	Theophylline	RNA	Escherichia coli	Growth selection	LacZ/CAT/GFP/TetA	Synthetic aptamer mTCT8-4 aptamer	RBS	[45,48]
	Theophylline	RNA	Saccharomyces cerevisiae	FACS	GFP	Synthetic aptamer mTCT8-4 aptamer	Hammerhead ribozyme	[50,46,47]

Tab.1 Different types of small-molecule biosensors and their applications in high-throughput screens^a)

Fig.3 Schematic representation of different biosensor design strategies as well as the coupled reporters and corresponding screening methods

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