微生物学报

Metal-responsive protein family	Family members and responders	References
MerR family	MerR (Hg), ZntR (Cd, Pb, Zn), ZccR (Zn, PbrR (Pb), CueR (Cu, Au), GolS (Au), HmrR (Ag), CoaR (Co), CadR (Cd), NimR (Ni)	[27-30]
Fur family	Fur (Fe), Zur (Zn), Mur (Mn), Nur (Ni), PerR (Mn/Fe)	[31-32]
DtxR family	DtxR, IdeR, SirR (Fe), MntR (Mn, ScaR (Cd), TroR (Mn, Zn)	[33-36]
NikR family	NikR (Ni)	[37-38]
ArsR/SmtB family	CmtR, ZiaR, AztR (Zn), CzrA (Zn, Co), ArsR (As, Sb, Bi), CadC (Cd, Pb, Zn), SmtB (As, Cd), NmtR (Ni, Co), KmtR (Ni), BxmR (Cu, Ag, Zn, Cd)	[39-44]
CsoR-RcnR family	CsoR (Cu), RcnR (Ni、Co)	[45-47]
CopY family	CopY (Cu)	[48]

Metal-responsive protein family	Family members and responders	References
MerR family	MerR (Hg), ZntR (Cd, Pb, Zn), ZccR (Zn, PbrR (Pb), CueR (Cu, Au), GolS (Au), HmrR (Ag), CoaR (Co), CadR (Cd), NimR (Ni)	[27-30]
Fur family	Fur (Fe), Zur (Zn), Mur (Mn), Nur (Ni), PerR (Mn/Fe)	[31-32]
DtxR family	DtxR, IdeR, SirR (Fe), MntR (Mn, ScaR (Cd), TroR (Mn, Zn)	[33-36]
NikR family	NikR (Ni)	[37-38]
ArsR/SmtB family	CmtR, ZiaR, AztR (Zn), CzrA (Zn, Co), ArsR (As, Sb, Bi), CadC (Cd, Pb, Zn), SmtB (As, Cd), NmtR (Ni, Co), KmtR (Ni), BxmR (Cu, Ag, Zn, Cd)	[39-44]
CsoR-RcnR family	CsoR (Cu), RcnR (Ni、Co)	[45-47]
CopY family	CopY (Cu)	[48]

Sensing element	Detection object	Reporter gene	Host cell	Application	Detection range (μmol/L)	References
Transcription activator
MerR	Hg²⁺	rfp	Escherichia coli DH5α	Rapid and convenient screening of total inorganic mercury in cosmetics	0.050 0-10	[60]
	Hg²⁺	rfp, amilcpblue	Escherichia coli DH5α	The extremely wide linear range meets the different monitoring requirements	0.001 0-1 (Mer-RFP), 0.002 0-0.125 0 (Mer-Blue)	[61]
ZntR	Zn²⁺	zntR, ribB, oprF	Escherichia coli BL21	Microbial fuel cell electrobiosensor	0-400	[62]
PbrR	Pb²⁺	vio ABCDE	Escherichia coli TOP10	Detection of biologically available Pb²⁺ in water	0.187 5-1.500 0	[63]
	Pb²⁺	Luc	Escherichia coli DH5α	Detection of Pb²⁺ in environmental and biological samples	1-100	[64]
CadR	Cd²⁺	vio ABCDE	Escherichia coli TOP10	Detection of soluble Cd²⁺ in environmental water samples	0.049 0-25	[65]
	Cd²⁺	rfp	Multiple Gram-negative bacteria	The importance of WCMB testing in different genera of bacteria was emphasized	0.500 0–2.000 0 μg/mL (E. coli DH5α) 0.100 0 μg/mL (Pseudomonas aeruginosa PAO1) 10 μg/mL (Shewanella oneidensis MR-1) 0.250 0 μg/mL (Enterobacter sp. NCR3) 1 μg/mL (Enterobacter sp. LCR17)	[66]
	Pb²⁺	Luc	Escherichia coli DH5α	For the detection of Pb²⁺ in environmental and biological samples	0.010 0-10	[64]
CueR	Au³⁺	rfp	Cupriavidus etallidurans	Monitoring the concentration of Au³⁺ in human urine	≥0.110 0	[67]
Transcription inhibitors
EcArsR	AsO₃^3-	luc	Escherichia coli DH5α	A WCMB that is highly sensitive to arsenite has been developed	0.100 0-1≥0.040 0	[68]
ArsR1	AsO₃^3-	gfp	Escherichia coli TOP10	A WCMB that is highly sensitive to arsenite has been developed	0.030 0-0.100 0 (2.250 0-7.500 0 μg/L) ≥0.010 0	[69]
SxArsR	Sb³⁺	luc	Sphingobium xenophagum C1	A novel subtype of the ArsR family transcription factor, designated as SxArsR, has been identified, which exhibits specific responsiveness to Sb	0.010 0-6 ≥0.010 0^*	[70]
Combination of transcription factors
MerR, CadR	Hg²⁺, Cd²⁺	egfp, mCherry	Escherichia coli TOP10	Detection of concurrent heavy metal contaminants in the environment	0-40 (Hg²⁺), 0-200 (Cd²⁺)	[71]
CadC, CadR	Cd²⁺	egfp and mCherry	Escherichia coli TOP10	Detection of high concentrations of biologically available Cd in water	≥0.050 0 (CadC-eGFP), ≥0.100 0 (CadR-mCherry)	[72]
MerR, PbrR	Hg²⁺, Pb²⁺	Indigoidine biosynthetic module	Escherichia coli TOP10	Detection of Hg²⁺ and Pb²⁺ in environmental samples	≥0.008 0	[73]
CadR, MerR	Cd²⁺, Pb²⁺, Hg²⁺	vioABE, vioC	Escherichia coli TOP10	Designed for detecting heavy metal contaminants in seawater	0.004 9-40, ≥0.004 9 (Cd²⁺), 0.024 4-200, ≥0.024 4 (Pb²⁺), 0.003 7-0.468 8, ≥0.000 5 (Hg²⁺)^*	[74]
UzcRS, UrpRS	UO₂²⁺	gfp	Caulobacter vibrioides	Detection of uranium (U) in groundwater samples	≥1	[75-76]

Sensing element	Detection object	Reporter gene	Host cell	Application	Detection range (μmol/L)	References
Transcription activator
MerR	Hg²⁺	rfp	Escherichia coli DH5α	Rapid and convenient screening of total inorganic mercury in cosmetics	0.050 0-10	[60]
	Hg²⁺	rfp, amilcpblue	Escherichia coli DH5α	The extremely wide linear range meets the different monitoring requirements	0.001 0-1 (Mer-RFP), 0.002 0-0.125 0 (Mer-Blue)	[61]
ZntR	Zn²⁺	zntR, ribB, oprF	Escherichia coli BL21	Microbial fuel cell electrobiosensor	0-400	[62]
PbrR	Pb²⁺	vio ABCDE	Escherichia coli TOP10	Detection of biologically available Pb²⁺ in water	0.187 5-1.500 0	[63]
	Pb²⁺	Luc	Escherichia coli DH5α	Detection of Pb²⁺ in environmental and biological samples	1-100	[64]
CadR	Cd²⁺	vio ABCDE	Escherichia coli TOP10	Detection of soluble Cd²⁺ in environmental water samples	0.049 0-25	[65]
	Cd²⁺	rfp	Multiple Gram-negative bacteria	The importance of WCMB testing in different genera of bacteria was emphasized	0.500 0–2.000 0 μg/mL (E. coli DH5α) 0.100 0 μg/mL (Pseudomonas aeruginosa PAO1) 10 μg/mL (Shewanella oneidensis MR-1) 0.250 0 μg/mL (Enterobacter sp. NCR3) 1 μg/mL (Enterobacter sp. LCR17)	[66]
	Pb²⁺	Luc	Escherichia coli DH5α	For the detection of Pb²⁺ in environmental and biological samples	0.010 0-10	[64]
CueR	Au³⁺	rfp	Cupriavidus etallidurans	Monitoring the concentration of Au³⁺ in human urine	≥0.110 0	[67]
Transcription inhibitors
EcArsR	AsO₃^3-	luc	Escherichia coli DH5α	A WCMB that is highly sensitive to arsenite has been developed	0.100 0-1≥0.040 0	[68]
ArsR1	AsO₃^3-	gfp	Escherichia coli TOP10	A WCMB that is highly sensitive to arsenite has been developed	0.030 0-0.100 0 (2.250 0-7.500 0 μg/L) ≥0.010 0	[69]
SxArsR	Sb³⁺	luc	Sphingobium xenophagum C1	A novel subtype of the ArsR family transcription factor, designated as SxArsR, has been identified, which exhibits specific responsiveness to Sb	0.010 0-6 ≥0.010 0^*	[70]
Combination of transcription factors
MerR, CadR	Hg²⁺, Cd²⁺	egfp, mCherry	Escherichia coli TOP10	Detection of concurrent heavy metal contaminants in the environment	0-40 (Hg²⁺), 0-200 (Cd²⁺)	[71]
CadC, CadR	Cd²⁺	egfp and mCherry	Escherichia coli TOP10	Detection of high concentrations of biologically available Cd in water	≥0.050 0 (CadC-eGFP), ≥0.100 0 (CadR-mCherry)	[72]
MerR, PbrR	Hg²⁺, Pb²⁺	Indigoidine biosynthetic module	Escherichia coli TOP10	Detection of Hg²⁺ and Pb²⁺ in environmental samples	≥0.008 0	[73]
CadR, MerR	Cd²⁺, Pb²⁺, Hg²⁺	vioABE, vioC	Escherichia coli TOP10	Designed for detecting heavy metal contaminants in seawater	0.004 9-40, ≥0.004 9 (Cd²⁺), 0.024 4-200, ≥0.024 4 (Pb²⁺), 0.003 7-0.468 8, ≥0.000 5 (Hg²⁺)^*	[74]
UzcRS, UrpRS	UO₂²⁺	gfp	Caulobacter vibrioides	Detection of uranium (U) in groundwater samples	≥1	[75-76]

Sensing element	Detection object	Optimization strategy	Application	Performance enhancement (μmol/L)	References
ZntR	Cr²⁺ or Pb²⁺	Protein engineering	Cr and Pb-responsive WCMB were developed from the znt-manipulation subsystem for the determination of PB²⁺ or Cr²⁺ in environmental system	New selectivity for Cr²⁺ or Pb²⁺	[90]
	Pb²⁺	Protein engineering	The content of Pb²⁺ in environmental samples was monitored	0-0.010 0, <0.005 0^*	[58]
ArsR	Pb²⁺	Protein engineering	The content of Pb²⁺ in environmental samples was monitored	Detection accuracy>95%	[91]
MerR	Hg²⁺	Protein engineering	The content of Hg²⁺ in water samples was monitored	≥0.020 4	[92]
	Hg²⁺ Zn²⁺ Cu²⁺	Protein engineering	A new chimeric regulator WCMB was developed by replacing the metal-binding domain of MerR with ZntR or CueR	≥0.001 0 Hg²⁺ (MerR-Luc) ≥30 Zn²⁺ (MerRZntR-Luc) ≥10 Cu²⁺ (MerRCueR-Luc)	[93]
CadR	Cd²⁺	Protein engineering	The content of Cd²⁺ in water samples was monitored	≥0.450 0 μg/L	[94]
	Cd²⁺	Protein engineering	The interference of Hg²⁺ was reduced and the response to Cd²⁺ was enhanced	0.500 0-100， ≥0.079 0^*	[30]
MerR	Hg²⁺	Protein engineering	The “Parabola principle” is proposed and a visualized WCMB for Hg²⁺ detection in natural water is developed	0.200 0-0.250 0	[95]
	Hg²⁺	Protein engineering Promoter engineering	A super-sensitive visualized WCMB for the detection of ultra-trace Hg²⁺ was developed	≥0.313 0ng/L. When the fluorescence signal is more than 2.500 0 ng/l, it can be observed directly^*	[96]
ArsR	AsO₃^3-	Promoter engineering	A high sensitivity AsO₃^3- WCMB was developed, and the background reduction and signal output improvement were realized	0.100 0-4, ≥0.010 0	[97]
	AsO₃^3-	Promoter engineering	A highly sensitive and specific WCMB for AsO₃^3- detection in drinking water was developed	≥0.010 0^*	[98]
	AsO₃^3-	Promoter engineering	A functional promoter library screening method was developed, and different AsO₃^3- responsive elements arsR and OsmE1 were obtained, and a novel OsmE1 biosensor was constructed	≥0.040 0	[99]
	AsO₃^3-	Promoter engineering	A highly sensitive and specific WCMB for AsO₃^3- detection in drinking water was developed	≥0.100 0	[100]
ArsR MerR	AsO₃^3- Hg²⁺	Promoter engineering	A modular cascaded signal amplification method was developed, and a low-cost, portable and accurate AsO₃^3- and Hg²⁺ ultra-sensitive WCMB was constructed	≥0.100 0 ppb AsO₃^3- ≥0.010 0 ppb Hg²⁺	[101]
CadR	Cd²⁺	Promoter engineering	A highly sensitive and specific Cd²⁺ responsive WCMB was developed for the detection of Cd²⁺ in water	≥0.010 0	[102]
	Cd²⁺	Promoter engineering	A Cd²⁺ responsive WCMB with a negative feedback amplifier was developed to detect Cd²⁺ below the WHO and FAO standards with extremely high sensitivity and specificity	≥0.000 1^*	[103]

Sensing element	Detection object	Optimization strategy	Application	Performance enhancement (μmol/L)	References
ZntR	Cr²⁺ or Pb²⁺	Protein engineering	Cr and Pb-responsive WCMB were developed from the znt-manipulation subsystem for the determination of PB²⁺ or Cr²⁺ in environmental system	New selectivity for Cr²⁺ or Pb²⁺	[90]
	Pb²⁺	Protein engineering	The content of Pb²⁺ in environmental samples was monitored	0-0.010 0, <0.005 0^*	[58]
ArsR	Pb²⁺	Protein engineering	The content of Pb²⁺ in environmental samples was monitored	Detection accuracy>95%	[91]
MerR	Hg²⁺	Protein engineering	The content of Hg²⁺ in water samples was monitored	≥0.020 4	[92]
	Hg²⁺ Zn²⁺ Cu²⁺	Protein engineering	A new chimeric regulator WCMB was developed by replacing the metal-binding domain of MerR with ZntR or CueR	≥0.001 0 Hg²⁺ (MerR-Luc) ≥30 Zn²⁺ (MerRZntR-Luc) ≥10 Cu²⁺ (MerRCueR-Luc)	[93]
CadR	Cd²⁺	Protein engineering	The content of Cd²⁺ in water samples was monitored	≥0.450 0 μg/L	[94]
	Cd²⁺	Protein engineering	The interference of Hg²⁺ was reduced and the response to Cd²⁺ was enhanced	0.500 0-100， ≥0.079 0^*	[30]
MerR	Hg²⁺	Protein engineering	The “Parabola principle” is proposed and a visualized WCMB for Hg²⁺ detection in natural water is developed	0.200 0-0.250 0	[95]
	Hg²⁺	Protein engineering Promoter engineering	A super-sensitive visualized WCMB for the detection of ultra-trace Hg²⁺ was developed	≥0.313 0ng/L. When the fluorescence signal is more than 2.500 0 ng/l, it can be observed directly^*	[96]
ArsR	AsO₃^3-	Promoter engineering	A high sensitivity AsO₃^3- WCMB was developed, and the background reduction and signal output improvement were realized	0.100 0-4, ≥0.010 0	[97]
	AsO₃^3-	Promoter engineering	A highly sensitive and specific WCMB for AsO₃^3- detection in drinking water was developed	≥0.010 0^*	[98]
	AsO₃^3-	Promoter engineering	A functional promoter library screening method was developed, and different AsO₃^3- responsive elements arsR and OsmE1 were obtained, and a novel OsmE1 biosensor was constructed	≥0.040 0	[99]
	AsO₃^3-	Promoter engineering	A highly sensitive and specific WCMB for AsO₃^3- detection in drinking water was developed	≥0.100 0	[100]
ArsR MerR	AsO₃^3- Hg²⁺	Promoter engineering	A modular cascaded signal amplification method was developed, and a low-cost, portable and accurate AsO₃^3- and Hg²⁺ ultra-sensitive WCMB was constructed	≥0.100 0 ppb AsO₃^3- ≥0.010 0 ppb Hg²⁺	[101]
CadR	Cd²⁺	Promoter engineering	A highly sensitive and specific Cd²⁺ responsive WCMB was developed for the detection of Cd²⁺ in water	≥0.010 0	[102]
	Cd²⁺	Promoter engineering	A Cd²⁺ responsive WCMB with a negative feedback amplifier was developed to detect Cd²⁺ below the WHO and FAO standards with extremely high sensitivity and specificity	≥0.000 1^*	[103]

重金属全细胞微生物传感器：设计原理、优化策略及监测应用

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王明月 ¹^,² , 许玫英 ² , 纠敏 ¹^,^* , 陈杏娟 ²^,^*

微生物学报 | 综述 2025,65(3): 1033-1052

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微生物学报 | 综述 2025, 65(3): 1033-1052

重金属全细胞微生物传感器：设计原理、优化策略及监测应用

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王明月¹^,², 许玫英², 纠敏¹^,^*, 陈杏娟²^,^*

作者信息

1 河南科技大学食品与生物工程学院，河南洛阳

2 广东省科学院微生物研究所，华南应用微生物国家重点实验室，广东省菌种保藏与应用重点实验室，广东广州

Whole-cell microbial biosensors for heavy metals: design principles, optimization strategies, and monitoring applications

Mingyue WANG¹^,², Meiying XU², Min JIU¹^,^*, Xingjuan CHEN²^,^*

Affiliations

1 College of Food and Bioengineering, Henan University of Science and Technology, Luoyang, Henan, China

2 State Key Laboratory of Applied Microbiology Southern China, Guangdong Provincial Key Laboratory of Microbial Culture Collection and Application, Institute of Microbiology, Guangdong Academy of Sciences, Guangzhou, Guangdong, China

出版时间: 2025-03-04 doi: 10.13343/j.cnki.wsxb.20240619

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摘要

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重金属因其毒性、持久性、生物蓄积性以及对抗性基因的潜在贡献等特点，已引起人们越来越多的关注。对环境中重金属进行准确、快速、高效和灵敏的监测，对于保护环境和人体健康具有重要意义。全细胞微生物传感器作为一种集成生物识别模块与传感处理模块的生物监测技术，为环境重金属污染的监测提供了新的解决方案。近年来，基于转录因子的全细胞微生物传感器在重金属监测应用方面取得了显著进展。本文综述了重金属全细胞微生物传感器的基本组成及设计原理，总结了近年来研发的重金属全细胞微生物传感器的构建及其应用情况，分析了通过合成生物学技术优化重金属全细胞微生物传感器检测性能的应用实例，并展望了该领域面临的挑战及未来可能的研究方向。本文有望为环境中重金属污染的高效监测与有效防控提供重要参考。

关键词

重金属 / 全细胞微生物传感器 / 设计 / 优化 / 应用

Abstract

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Heavy metal pollution has attracted increasing attention because of its toxicity, environmental persistence, bioaccumulation, and potential contribution to antibiotic resistance. The accurate, rapid, efficient, and sensitive detection of heavy metals in the environment is of great significance for environmental protection and human health. A whole-cell microbial biosensor (WCMB), integrating a biorecognition module and a signal processing module, provides a new strategy for heavy metal monitoring. Great progress has been achieved in the transcription factor-based WCMBs for the monitoring of heavy metals in recent years. This paper introduced the basic composition and design principles of WCMBs, summarized the construction and application of WCMBs for heavy metal detection that were developed in recent years, analyzed the optimization strategies of WCMBs by synthetic biology, and finally prospected the future research directions of WCMBs. This paper is expected to be an important reference for the effective prevention and control of heavy metal pollution in the environment.

Key words

heavy metals / whole-cell microbial biosensor / design / optimization / application

引用本文

王明月, 许玫英, 纠敏, 陈杏娟. 重金属全细胞微生物传感器：设计原理、优化策略及监测应用. 微生物学报, 2025 , 65 (3) : 1033 -1052 . DOI: 10.13343/j.cnki.wsxb.20240619

Mingyue WANG, Meiying XU, Min JIU, Xingjuan CHEN. Whole-cell microbial biosensors for heavy metals: design principles, optimization strategies, and monitoring applications[J]. Acta Microbiologica Sinica, 2025 , 65 (3) : 1033 -1052 . DOI: 10.13343/j.cnki.wsxb.20240619

正文

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重金属一般是指密度大于5 g/cm³和分子量大于40.04的金属元素^[1]。它们主要通过自然过程或人类活动进入环境，并以各种形式在食物链中富集^[2-3]。重金属在人体内蓄积会引起多种健康问题，如中枢神经毒性、生殖毒性、心血管疾病甚至癌症等^[4]；在水生生物体内蓄积会影响鱼虾等的生长和繁殖，最终可能导致整个生态系统的失衡和破坏^[5]。重金属在土壤中积累会导致土壤酸化、结构破坏和肥力下降，影响农作物的生长发育^[6]。农产品和食品中的重金属污染更会引发严重的健康和食用安全风险^[7]。此外，越来越多的研究表明，环境中的重金属污染还会加剧细菌的抗生素耐药性，构成更为广泛的全球健康威胁^[8]。因此，环境中的重金属污染问题已引起了人们越来越多的关注^[9]。

世界各国和组织均高度重视重金属污染控制问题，针对不同类型的重金属均制定了相应的限制标准和管制措施^[10-11]。这些不断更新的限量标准和法规给重金属检测和污染控制带来了新的挑战。目前已有多种化学分析方法用于重金属的检测，例如电感耦合等离子体质谱法、原子荧光光谱法等；这些方法灵敏度高，识别特异性强^[12]。然而，它们往往需要昂贵的仪器设备以及专业的测量人员，样品制备过程繁琐，并且无法提供生物可利用度及生物毒性效应等相关信息；为了克服这些限制，人们开始关注以酶、抗体和微生物细胞等生物材料作为识别模块的生物传感器研究^[13]。

生物传感器的基本工作原理主要涉及生物识别与信号传感(图1A)。生物识别主要通过高选择性和高灵敏性的生物受体(如全细胞、酶、抗体和DNA等)与检测物直接接触而发挥作用，该过程会产生各种报告信号变化，如发光、产热、产生电子、pH改变以及颜色生成等^[14]。信号传感则主要通过传感处理模块将报告信号变化转换为可被检测的形式；生物传感器具有快捷简便、成本低廉、高效灵敏等优点，不仅省去了繁琐的前处理操作，还具有生物毒性效应预警等功能^[15]。目前，生物传感器已广泛应用于临床诊断、环境监测、食品安全和地质勘探等领域^[16-19]。

全细胞微生物传感器(whole-cell microbial biosensor, WCMB)采用完整的微生物细胞集成生物识别模块与信号传感模块，与酶、核酸生物传感器相比，其制备成本更低、操作性能更稳定，能够在复杂环境中生存和适应，并可重复使用；其中以转录因子作为生物识别模块构建的重金属WCMB数量最多、应用最广，不仅展现出对多种重金属离子的高灵敏性和特异性响应，还具备简单快速、现场检测的能力^[20]。

虽然早在10年前已有研究者对重金属WCMB的信号输出/输入、特异性/耐受性、底盘细胞选择以及环境应用、安全风险及监管等方面进行了详细综述^[21]，但当时重金属WCMB的特异性及灵敏性仍有较大提升空间。近10年来，随着合成生物学技术方法的飞速发展，基于转录因子的重金属WCMB的特异性与灵敏性得到了前所未有的提升，其实际环境应用也得到了极大的拓展。本文在前人研究的基础上，更新了大量研究数据和成果，进一步探讨了WCMB在重金属监测领域的最新进展和未来趋势。本文系统阐述了基于转录因子的重金属WCMB的基本组成及设计原理，总结了近年来研发的重金属WCMB的构建策略及应用，分析了通过合成生物学技术方法优化WCMB检测性能的应用实例，并展望了该领域面临的挑战和未来可能的研究方向，以期为环境中重金属污染的高效监测与有效防控提供重要参考。

1 基于转录因子的重金属全细胞微生物传感器的构建及应用

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1.1 全细胞微生物传感器的基本组成和设计原理

WCMB以完整细胞集成生物识别模块与传感处理模块^[20]，包含响应元件及其结合的启动子及响应元件触发的下游信号报告元件，其对重金属的识别与传感过程如图1B所示。WCMB的设计是一个复杂的过程，主要包括响应元件选择、基因回路设计、工程细胞构建、环境测试以及性能优化等步骤^[22]。

1.1.1 响应元件的选择

响应元件通常由一组响应重金属的转录因子及其启动子组成；基于转录因子构建的WCMB数量最多、应用最广，其工作原理模仿微生物的重金属抗性系统，依赖重金属与转录因子相互作用，从而驱动报告信号基因表达^[23]。当环境中的重金属穿过细胞膜并与特定的转录因子相互作用时，转录因子的蛋白构象会发生变化，进而影响转录因子与基因表达调控区域之间的空间位阻或结合亲和力，从而使转录因子结合特定启动子以激活报告基因表达，或者从启动子区域释放以解除阻遏并启动报告基因表达^[24](图2)。报告基因的转录和翻译会产生能被检测的信号；这种报告信号受转录因子调控，而转录因子构象的改变通常与重金属浓度相关，使生物传感器具有感应不同浓度重金属的能力^[25]。金属抗性系统通常包括金属响应调控蛋白(转录因子)、金属转运蛋白、金属结合蛋白等^[26]。目前，已知的金属响应调控蛋白家族主要有7类，如表1所示。

1.1.2 报告元件的选择

生物发光或荧光等光信号传导方式，因其敏感性及报告基因的多样性而备受青睐。这些报告基因可以是荧光蛋白或荧光素酶；常用的荧光蛋白包括绿色荧光蛋白(green fluorescent protein, GFP)、黄色荧光蛋白(yellow fluorescent protein, YFP)、红色荧光蛋白(red fluorescent protein, RFP)等；这些蛋白在特定波长的光激发下能够发出稳定且明亮的荧光，对细胞无毒害且易于检测^[49]。荧光素酶能够催化生物体产生荧光，具有高灵敏度和可定量检测等特点，因此也是理想的报告基因^[50]。此外，产酸碱蛋白、色素合成蛋白、电子介体蛋白等也是常见的生物传感器报告元件^[51]。

1.1.3 基因回路的设计

基因回路主要由调节元件和被调节元件组成；调节元件通常包括启动子和转录因子等，它们负责重金属的识别和转录调控；被调节元件则多为编码信号蛋白的基因，负责产生输出信号；金属WCMB基因回路的设计可以有多种方案，如图3所示^[52]，最常见的通路有4种：(1)单输入信号通路；(2)双输入单诱导信号通路，报告信号由1个输入诱导；(3)双输入双诱导信号通路，报告信号由2个输入同时诱导；(4)多输入信号通路，用于开发需要复杂调控系统的多输入传感器。通过合理设计基因回路可以改善生物传感器的各种参数。

1.1.4 底盘细胞的选择

底盘细胞作为监测重金属浓度及毒性的受体宿主，应具备良好的环境适应能力和遗传稳定性，以提升传感器的可靠性、重复性和稳定性；此外，底盘细胞对目标检测物应具有较高的敏感性，能在低目标物浓度下迅速响应，从而拓宽检测范围并提高精度；底盘细胞还应能够特异性地识别目标检测物，避免交叉反应，减少干扰和误差，进而提高传感器的准确性^[53]。在实际应用中，常用的底盘细胞包括细菌、酵母、藻类、原生动物甚至植物细胞等^[54-57]。细菌因生长迅速且环境适应能力强，与其他真核生物相比被使用的频率更高；随着基因工程与分子生物学等领域的快速发展，经过改造的大肠杆菌(Escherichia coli)和其他类型的基因工程菌也被广泛应用于WCMB的构建中^[58]。

1.2 基于转录因子的重金属全细胞微生物传感器的构建及应用

转录因子是生物传感器中最常见的响应元件，能够识别并结合特定的DNA序列，从而调控相关基因的表达。由于转录因子具有结合特异性和信号转导能力，因此转录因子的选择与设计决定了WCMB的检测性能。转录因子通常由DNA结合域和配体结合域组成；根据其作用机制可分为转录激活因子和转录抑制因子；根据变构方式又可分为单组分转录因子和双组分转录因子^[59]。近年来，基于不同类型转录因子构建的生物传感器在环境重金属检测中的应用实例见表2。

1.2.1 基于转录激活因子的重金属全细胞微生物传感器的构建及应用

MerR、ZntR、PbrR、CadR、CueR、CupR以及GolS是细菌中用于重金属解毒的转录激活因子^[27-30]。利用这些转录激活因子与相应重金属结合后促使报告基因表达的原理，研究者们开发了多种监测Hg²⁺、Zn²⁺、Pb²⁺、Cd²⁺、Au³⁺等重金属的全细胞生物传感器，并成功应用于化妆品、饮用水和尿液等环境样品中的重金属监测，为环境污染监测与人体健康监测提供了新的有力工具^{[60-63,65-67,77-78]}。

MerR家族的金属调控蛋白是一种被广泛用于构建WCMB的关键元件，通常具有保守的N端DNA结合域和结构存在差异的C端金属结合域，能够特异性地与金属离子结合，导致蛋白构象扭曲，从而容易被RNA聚合酶识别并转录激活下游报告基因的表达，产生可被检测的信号输出^[77]。以MerR作为特异性识别元件及RFP作为报告元件，在E. coli DH5α中构建的汞响应WCMB对0.050 0-10 μmol/L的Hg²⁺具有良好的线性响应能力(R²=0.984 8)；通过将该WCMB嵌入滤纸制成测试条，实现了对化妆品中总无机汞污染物的快速、便捷的现场筛查；其对Hg²⁺离子的检测下限为1 μmol/L，当Hg²⁺离子浓度达到5 μmol/L时，整个试纸会呈现出明显的红色；此外，该WCMB不仅能够响应Hg²⁺，还能够响应不溶性的Hg₂Cl₂和Hg(NH₂)Cl^[60]。同样，以MerR作为特异性识别元件，以及AmilCP和RFP蛋白作为报告元件，开发的2种汞响应WCMB (Mer-Blue和Mer-RFP)

对Hg²⁺具有特异性响应；其中Mer-Blue的线性范围为2-125 nmol/L，而Mer-RFP具有1-1 000 nmol/L的极宽线性范围，适用于不同环境条件下的监测需求^[61]。与MerR类似，MerR家族的另一个成员ZntR也被常用作WCMB的构建元件。以ZntR作为识别元件及核黄素合成蛋白RibB作为报告元件，在E. coli BL21中设计了受PzntA调控的zntR-ribB-oprF基因融合表达系统，并引入微生物燃料电池MFC设计，构建了锌响应电化学WCMB；该WCMB能够灵敏感知Zn²⁺并促使核黄素的产生及电压的输出，实现了Zn²⁺在0-400 μmol/L的线性检测(R²=0.977 7)^[62]，为重金属离子的检测提供了一种新的思路和方法。

铅响应转录调控因子PbrR的金属结合域能够特异性地结合Pb²⁺，使其在构建铅响应WCMB中具有重要意义；以PbrR蛋白作为识别元件及紫罗兰素生物合成蛋白VioABCDE作为报告元件，在E. coli TOP10中构建的铅响应WCMB可检测线性范围为0.187 5-1.500 0 μmol/L的Pb²⁺，实现了对Pb²⁺肉眼可见的定性检测，提高了检测的便捷性和直观性^[63]。

CadR金属调控蛋白具有Cys77、Cys112、Cys119等3个保守的金属结合位点，对Cd²⁺具有高度的特异性和亲和力^[78]。以CadR及其启动子作为识别模块，VioABCDE作为报告模块，构建了镉响应WCMB，对Cd²⁺的响应明显强于其他重金属离子(包括Pb²⁺、Zn²⁺和Hg²⁺)，可检测浓度低至0.049 0 μmol/L的Cd²⁺；当Cd²⁺浓度达到25 μmol/L时，通过肉眼即可观察到紫罗兰素颜色的变化^[65]。以红色荧光蛋白(RFP)作为报告元件，以CadR-GFP作为识别元件，在宽宿主范围的载体上进行组装并分别导入E. coli DH5α、铜绿假单胞菌(Pseudomonas aeruginosa) PAO1、奥奈达湖希瓦氏菌(Shewanella oneidensis) MR-1、肠杆菌属(Enterobacter)中，构建的不同宿主的镉响应WCMB (pBBcadRgfp-rfp)对Cd²⁺的线性范围和检测限存在显著差异；尽管条件相似，但多重镉响应WCMB可能受到所使用的宿主细胞种类的影响^[66]。

当以PbrR和CadR作为识别元件，荧光素酶蛋白Luc作为报告元件时构建了2种WCMB，其中pGL3-luc/cad可检测0.010 0-10 μmol/L的Pb²⁺，在浓度为300 μmol/L时才观察到铅对细胞的毒性；相比之下，pGL3-luc/pbr具有较高的灵敏度和特异性，可检测1-100 μmol/L的Pb²⁺，并且不受其他金属离子(如Sn²⁺、Ni²⁺、Cd²⁺)的干扰^[64]。

CueR蛋白作为转录激活因子，能够结合在启动子DNA的-35--10区之间，通过改变DNA构象来激活下游基因的转录；以CueR作为识别元件及红色荧光蛋白(RFP)作为报告元件，在耐重金属贪铜菌(Cupriavidus metallidurans)中构建的受启动子PcopZ驱动的金响应WCMB具有高度的Au³⁺响应灵敏度，最低检测限为110 nmol/L；基于智能手机分析系统，可以对金响应WCMB的红色荧光信号进行实时采集和分析，通过感知尿液样本中Au³⁺的存在来监测人体健康，降低了疾病诊断的检测成本并提高了检测便捷性^[67]。

尽管这些转录因子作为识别元件对重金属展现出高特异性，但在实际应用中，仍可能受到其他金属离子的影响，进而削弱WCMB的准确性和灵敏度。因此，持续的改进和优化工作仍然尤为重要。

1.2.2 基于转录抑制因子的重金属全细胞微生物传感器的构建及应用

目前，已有多种转录抑制因子被用于构建重金属监测WCMB，其中最典型的是来自细菌砷解毒系统中的砷结合蛋白ArsR；ArsR能够与ars操纵子的启动子序列结合，是ars基因簇的负调控因子；当ArsR与砷结合后从启动子区域释放，从而促使编码相关解毒蛋白的ars操纵子中的基因转录^[79]。

目前ArsR家族分为5种亚型，分别具有独特的As/Sb结合位点^[80]。I型ArsR以E. coli质粒R773中的EcArsR为代表，其砷结合域含有3个半胱氨酸残基的保守“CXCXXC”结构，分别位于Cys32、Cys34和Cys37位^[81]。II型ArsR以谷氨酸棒杆菌(Corynebacterium glutamicum)中的CgArsR为代表，其砷结合域具有保守的半胱氨酸残基“CCX(42-43)CXC”结构，形成结合位点的3个半胱氨酸中的Cys15和Cys16位于一个二聚体亚基，而第3个Cys55则位于另一个二聚体亚基^[82]。III型ArsR以亚铁氧化酸硫杆状菌(Acidithiobacillus ferrooxidans)中的AfArsR为代表，其砷结合域具有保守的半胱氨酸残基“CCX(4-7)C”结构，分别为Cys95、Cys96和Cys102，位于C端柔性末端^[83]。IV型ArsR以腐败希瓦氏菌(Shewanella putrefaciens)中的SpArsR为代表，其砷结合域具有由Cys83、Cys101和Cys102组成的保守半胱氨酸残基“CX(17)CC”结构，参与甲基AsO₃^3-的解毒而对AsO₃^3-无响应^[84]。V型ArsR以食异源物鞘氨醇菌(Sphingobium xenophagum)的SxArsR为代表，是Chen等^[70]发现的一种新型ArsR转录抑制因子，具有保守的“HCXC”结构域，分别位于His28、Cys29和Cys31位；与其他4种已表征的ArsR亚型相比，新型SxArsR缺少第3个与AsO₃^3-结合的半胱氨酸残基，是Sb₂O₃的特异性结合位点。

基于I型ArsR构建的砷响应WCMB是研究最多的一种，其中Fang等^[68]基于E. coli筛选和优化了EcArsR及其变构体作为识别元件，以荧光素酶作为报告元件，构建了一系列砷响应WCMB，实现了对水体中低浓度AsO₃^3- (0.100 0-1 µmol/L)的快速监测。以硫还原地杆菌(Geobacter sulfurreducens)的ArsR1蛋白作为识别元件并融合GFP作为报告元件构建的砷响应WCMB，在最佳测试条件下对AsO₃^3-检测限达到0.010 0 μmol/L，GFP荧光强度与AsO₃^3-浓度在0.030 0-0.100 0 μmol/L (2.250-7.500 μg/L)范围内呈线性相关^[69]。以III型AfArsR作为识别元件，替换GFP的第146-147位氨基酸并融合在其中，构建的荧光增强型砷响应WCMB对1 mmol/L的AsO₃^3-响应的荧光强度变化达到31.6%；将GFP的Met146和Gln263分别突变为Phe146和His263，并随机诱变Glu147-Pro148和Gly261-Asn262后，获得的突变体对AsO₃^3-响应的荧光强度变化最大达到44.9%^[85]。Chen等^[70]基于V型SxArsR作为识别元件并融合荧光素酶作为报告元件，构建了锑响应WCMB，在Sb₂O₃浓度为0.010 0-6 μmol/L范围内呈现极好的线性关系(R²=0.99)，检测限低至0.010 0 μmol/L；在实际应用中该新型WCMB对环境水样和沉积物样品中Sb₂O₃的检测也表现出良好的性能。

NikR是另一种被广泛用于构建重金属监测生物传感器的转录抑制因子。NikR是Ni依赖的转录调节蛋白，通过负调控Ni ABC型转运体(Nik ABCDE)的表达来应对过量的Ni摄入，在维持细菌细胞中的Ni稳态发挥关键作用^[86]。NikR最初以四聚体的形式存在，本身并不具备与DNA结合的能力，但Ni²⁺的结合使其能够识别特定的启动子序列，并抑制下游Ni²⁺摄取基因的表达；以E. coli nik操纵子的启动子及NikR作为响应元件并以eGFP作为报告元件，Yoon等^[87]构建了一种重金属响应WCMB；在对一系列重金属离子(如Ni²⁺、Cd²⁺、As³⁺、Hg²⁺、Cu²⁺、Cr⁵⁺、Pb²⁺、Zn²⁺)的监测中发现，只有As³⁺被该WCMB响应；通过对NikR的结构分析，发现其金属结合域中的一些残基(如His62、His79、His92等)可能有助于As³⁺的结合；As³⁺的结合导致NikR的构象变化，使其从nik操纵子的启动子区释放出来，从而激活egfp报告基因的转录。

1.2.3 基于组合转录因子的重金属全细胞微生物传感器的构建及应用

利用不同转录因子组合可以在单一传感细胞内构建多种传感模块，实现多种重金属的同时检测；目前已有多种基于转录因子组合的重金属WCMBs成功应用于实际环境中的重金属检测(表2)。以MerR和CadR作为独立的响应元件，分别控制eGFP和mCherry荧光蛋白的表达，构建了同时检测Hg²⁺和Cd²⁺的双通道WCMB；可通过双荧光信号检测0-40 μmol/L浓度范围的Hg²⁺，通过红色荧光信号检测浓度<200 μmol/L的Cd^2+[71]。此外，利用CadC和CadR作为响应元件，分别控制eGFP和mCherry荧光蛋白的表达，构建的镉响应WCMB能够分别通过CadC-eGFP系统产生绿色荧光和CadR-mCherry系统产生红色荧光检测低至0.050 0 μmol/L和0.100 0 μmol/L的Cd^2+[72]。利用MerR和PbrR作为响应元件，以靛蓝生物合成基因簇作为报告元件，分别构建的WCMB能够通过靛蓝比色法检测到低至0.008 0 μmol/L的Hg²⁺和低至0.008 0 μmol/L的Pb²⁺，在Hg²⁺和Pb²⁺浓度分别达到0.033 0 μmol/L和2.080 0 μmol/L时，可通过肉眼观察到靛蓝颜色变化，为现场快速检测超标重金属提供了便捷方法，实现了对水样中生物可利用Hg和Pb的检测^[73]。分别以CadR和MerR作为响应元件、以VioABE和VioC作为报告元件构建的一种新型双色WCMB，可检测低至4.900 0 nmol/L的Cd²⁺、24.400 0 nmol/L的Pb²⁺和0.500 0 nmol/L的Hg²⁺；并且在0.004 9-40 μmol/L的Cd²⁺和0.024 4-200 μmol/L的Pb²⁺浓度范围内观察到灰绿色强度增加，在0.003 7-0.468 8 μmol/L的Hg²⁺浓度范围内观察到紫色强度增加，并呈剂量依赖性^[74]。这种低成本、新型WCMB具有同时检测环境水样中多种有毒重金属的潜力。

双组分系统是细菌体内的另一种信号转导系统，由组氨酸激酶与应答受体/转录因子组成，分别负责信号识别和细胞行为调控；基于双组分转录因子构建的WCMB具有复杂的遗传调控网络，其在实际环境监测应用中的普及程度尚不及上述基于单组分转录因子系统构建的WCMB^[88]。铀(U)是一种放射性重金属元素，对细菌具有显著毒性；然而，细菌如何感知U的分子机制长久以来鲜有报道；Park等^[75]通过反向遗传策略筛选获得新月柄杆菌(Caulobacter vibrioides)中受UO₂²⁺强烈诱导表达的启动子PurcA的调控元件，揭示了一种新的双组分系统UzcRS。此外，他们还鉴定了另一种由UrpR和UrpS两个蛋白构成的U响应双组分系统UrpRS，通过集成这2种功能分离的UzcRS和UrpRS双组分系统，构建了铀响应WCMB，实现了对浓度低至1 μmol/L的UO₂²⁺的精准检测，为评估地下水等环境样本中的铀污染提供了有力工具^[76]。

2 基于合成生物学策略的全细胞微生物传感器性能改造与优化

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尽管现有的WCMB能够检测多种重金属，但仍面临着部分重金属因毒性机制复杂或环境浓度低而导致检测难度增大的挑战；此外，WCMB在灵敏度、特异性和稳定性方面仍有待提高，以满足更为复杂、精细的环境监测需求^[22]。针对这些挑战，利用快速发展的合成生物学技术方法对天然蛋白质及调控DNA序列进行特异性改造，能够极大地优化WCMB的传感性能；其中的两大核心策略，蛋白质工程通过精细设计与定向进化技术可以改造识别元件的结构与功能，启动子工程通过优化启动子与操纵子的调控序列可以改造报告元件的表达强度；对识别元件与报告元件的精准改造可以实现WCMB检测特异性与灵敏性的双重提升；这些策略的综合运用，不仅有助于深化人们对生物传感机制的理解，更为WCMB在实际应用中的性能优化开辟了新路径^[89]。近年来，合成生物学策略在WCMB性能优化与改造领域的具体应用实例见表3。

2.1 蛋白质工程——改变转录因子的蛋白结构

除了对特定重金属的专一性响应外，多数天然转录因子还对其他重金属具有非特异性响应，这在复杂环境应用中常导致多金属干扰问题；转录因子的特异性改造，对于解决实际应用过程中的响应干扰问题具有重要的意义^[22]。

基于已知转录因子的结构，利用人工智能模拟与预测蛋白质结构，通过精确改造关键氨基酸残基的位置、电荷及亲疏水性等，优化转录因子与目标分子的相互作用，从而设计出新型结构的转录因子^[58,90-91]。Kim等^[90]基于ZntR的晶体结构信息对ZntR的金属结合域 “CCGTAHSSVYCS” 的不同位点进行定点突变，构建了23个对重金属具有不同响应敏感性的传感细胞，并成功获得了对Cr²⁺或Pb²⁺具有更高选择性的WCMB，进一步拓展了基于ZntR构建的WCMB在重金属检测领域的应用范围；该研究将定点突变技术应用于ZntR的金属结合域中，改变了ZntR的金属结合特性，为开发具有更高特异性和敏感性的WCMB提供了可能。Jeon等^[58]不仅对ZntR金属结合域的不同位点(如C115I、C115S、T117del、H119R、C124S)进行了定点突变，还敲除了宿主细胞中的金属离子输出基因copA和zntA，所构建的WCMB实现了对Pb²⁺的特异性检测，在0-10 nmol/L的Pb²⁺浓度范围内呈线性相关，检测限低于5 nmol/L，足以监测食品中铅的最大允许值(1.200 0-6.000 0 µmol/L)。Lee等^[91]通过对ArsR的砷结合序列“ELCVCDLCTA”的不同位点进行精准改造，也构建了23个对重金属具有不同响应敏感度的突变体，实现了ArsR从As³⁺到Pb²⁺的识别功能的转换；同时通过敲除copA和zntA基因，减弱了WCMB对Cd²⁺和Zn²⁺的选择性，增强了WCMB对Pb²⁺的识别特异性。这些研究成果不仅改变了铅响应操纵子pbr是唯一用于铅响应WCMB构建的识别元件的传统观点，还为构建其他重金属WCMB提供了新的思路和方向。

除了定点突变以外，研究者们还通过DNA大片段的替换，改造出性能更为优越的WCMB。Mendoza等^[92]通过替换识别元件GolS的金属结合域为MerR蛋白的等效区域，不仅提升了原有WCMB对高浓度Hg²⁺的耐受性(高达5 μmol/L)，还保持了原有WCMB的高灵敏检测能力，检测限达到20.400 0 nmol/L。Ghataora等^[93]使用结构域交换策略设计并构建了对重金属离子高灵敏和高特异检测的WCMB，通过在枯草芽孢杆菌(Bacillus subtilis) TW2043中引入了E. coli的MerR回路，所构建的汞响应WCMB可检测低至1 nmol/L的Hg²⁺；随后对MerR与ZntR和CueR进行结构域交换，设计了具有特定金属选择性的嵌合体调节器(MerRZntR和MerRCueR)，结合Lux ABCDE蛋白构建了2种WCMB，其中嵌合体MerRZntR WCMB可检测低至0.030 0 mmol/L的Zn²⁺，MerRCueR WCMB可检测低至10 μmol/L的Cu²⁺，为设计具有新型特异性的金属检测传感器提供了可能。

此外，通过基因随机诱变与定向进化，可以筛选出与目标分子结合能力显著增强的突变体，从而提升转录因子的结合与调控性能^[30,94]。Cai等^[94]以CadR作为识别元件及GFP为报告元件构建的镉响应WCMB为原始模板，利用易错PCR在CadR蛋白编码区进行随机突变，经过多轮荧光激活细胞分选后，获得了一株性能更高的细菌突变体epCadR5，其对Cd²⁺的检测灵敏度比天然CadR构建的WCMB提高了6.8倍，检测限达到0.450 0 μg/L。Guo等^[30]通过随机突变方法改变了CadR金属结合位点氨基酸的空间位置，构建了以CadR为响应元件、以RFP为报告元件的镉响应ebCadR-RFP WCMB，优化后的WCMB降低了Hg²⁺的干扰并增强了对Cd²⁺的响应，线性动态浓度范围为0.500 0-100 μmol/L，检测限为0.079 0 μmol/L。

2.2 启动子工程——优化结合位点序列、增强报告信号

2.2.1 优化转录因子结合位点及RNA聚合酶结合位点序列

当了解目标基因的启动子序列及转录因子结合位点信息时，可通过优化启动子与操纵子的调控序列以增强报告元件的表达强度，这对提升WCMB的检测灵敏性具有重要作用。传统重金属WCMB的设计主要依赖于模拟细菌的重金属抗性系统，而Guo等^[95]设计了一系列不同强度的组成型启动子来表达CadR蛋白，以GFP作为报告元件构建了一系列WCMB，结果发现传感器蛋白的表达水平可以作为WCMB检测灵敏度的“调节器”，并且检测曲线的斜率与传感器蛋白的表达水平(启动子表达效率的对数值)之间的关系可以用抛物线曲线(R²=0.886 7)来表示，于是提出了“启动子-斜率的抛物线原理”；同时还验证了抛物线原理同样适用于基于MerR家族其他传感器蛋白(如CueR、PbrR)的WCMB，为MerR家族传感蛋白的WCMB设计提供了一种新的策略，即通过选择不同表达强度的启动子调整传感蛋白的表达水平来精确控制WCMB的检测灵敏度；基于此原理，选择了5个merR基因的启动子来表达MerR蛋白，构建了用于天然水体中Hg²⁺检测的可视化WCMB，其中P429-merR WCMB表现出较好的抗干扰能力，在Hg²⁺浓度为0.200 0-0.250 0 μmol/L时呈现出肉眼可见的绿色变化。

Chen等^[97]通过对arsR-ParsWT操纵子的-10位点进行一系列的突变，构建了7个启动子突变体的砷响应WCMB，为了减少突变体中的背景无诱导表达，在砷响应启动子-10位点的下游添加了第2个ArsR结合位点，实现了WCMB性能的显著提升；其中，启动子突变体ParsD-ABS-8的灵敏度是天然启动子的11倍，在0.100 0-4.000 0 μmol/L AsO₃^3-范围内显示出优异的剂量响应(R²=0.992 8)，检测限约10 nmol/L。通过研究砷响应蛋白ArsR基因的调控特征，Chen等^[98]发现了砷结合蛋白ArsR的DNA非保守碱基对在蛋白质-DNA结合和基因转录调控中发挥重要作用，DNA非保守碱基对可以改变ArsR蛋白与其靶DNA序列的结合，非保守碱基对的变化也可以影响ArsR蛋白与DNA结合后的功能；通过对非保守碱基对的定点突变改造，开发了更灵敏和准确的砷响应WCMB，对AsO₃^3-的检测限提高到了0.010 0 μmol/L，远低于世界卫生组织(World Health Organization, WHO)对饮用水中AsO₃^3-的限量标准(0.010 mg/L或0.130 0 μmol/L)，为饮用水安全监测提供了有力工具。此外，Li等^[99]利用一种新的响应启动子高通量筛选技术直接从基因组中挖掘重金属响应启动子片段，构建了功能性启动子文库，并成功获得了AsO₃^3-响应的不同组件，用于砷响应WCMB的构建，这种方法构建的功能性启动子文库可以有效地筛选和发现重金属胁迫刺激下的响应性启动子，不仅高效而且不依赖现有基因组和转录因子信息，为重金属WCMB的构建与优化开辟了新路径。

2.2.2 设计增强报告信号的基因回路

在复杂环境中，目标信号往往微弱且易受干扰；对此，可以通过对基因回路的巧妙设计，包括正反馈和负反馈机制等，来放大报告基因的响应强度，以显著提高检测的灵敏度和准确性，对增加生物传感器的检测范围具有重要的作用^[99]。

Jia等^[102]将T7 RNA聚合酶基因元件置于CadR转录因子及其启动子的调控下，随后串联T7 RNA聚合酶启动子以调控报告蛋白mCherry元件的表达，并且在mCherry元件上游添加了受CadR抑制的操纵元件cadO，使用该策略构建的镉响应WCMB对Cd²⁺的检测限可以低至0.010 0 μmol/L，并且具有优异的响应特异性；此外，他们还将群体感应因子受体蛋白LuxR元件置于ArsR转录因子及其启动子的调控之下，同时将报告蛋白mCherry元件及另一个拷贝的群体感应因子受体蛋白LuxR元件置于群体感应因子合成酶的启动子调控下，构建了2种基因调控回路，使得报告蛋白mCherry元件的表达受到正反馈调节，通过该策略构建的砷响应WCMB的检测灵敏度和特异性大大提高，其灵敏度比未经改造的WCMB提高了约10倍，对AsO₃^3-的检测限低至0.100 0 μmol/L^[100]。除此之外，负反馈基因回路改造也是提高WCMB灵敏性的一种常见方法。Zhang等^[103]将四环素阻遏蛋白TetR元件置于CadR转录因子的下游，使其显著抑制CadR启动子Plteto1的本底表达，从而大大降低了无Cd²⁺条件下的背景信号干扰；通过此策略改造的镉响应WCMB对Cd²⁺的灵敏度比未经改造的WCMB提高了400倍，检测限低至0.100 0 nmol/L，且对Cd²⁺的耐受性也显著增加。

在需要快速、大量检测样本的情况下，通过调整识别元件和报告元件的数量以及增加信号放大模块是一种更为有效地提高WCMB灵敏性的方法；通过简单的组件调整和数量增加，即可实现对信号的放大和检测灵敏度的提升，相较于复杂的基因回路设计，该策略在成本控制方面可能更具优势^[104]。Zhu等^[96]首先对汞响应转录激活因子MerR进行定向进化，获得了高灵敏度的突变体，其中突变体m4-1对Hg²⁺检测限低至0.313 0 ng/L；然后在m4-1的GFP报告元件下游通过优化的5′非编码区序列集成了另一个GFP报告模块，构建了荧光信号增强且可视化的WCMB；在Hg²⁺浓度低至2.500 0 ng/L时，荧光信号仍然可通过肉眼直接观测，实现了对超痕量Hg²⁺的灵敏检测。Wan等^[101]进一步探索了级联模块化的信号放大策略，通过调节细胞内ArsR识别元件的数量来提高AsO₃^3-的检测灵敏度，然后集成3个转录放大器，逐级提升GFP报告元件的表达水平；成功开发了针对AsO₃^3-和Hg²⁺检测的超灵敏WCMB，灵敏度分别提高了750倍和200倍，检测限分别低至0.100 0 ppb和0.010 0 ppb。在实际应用中，应根据具体需求和条件选择合适的优化策略或进行综合运用。目前，通过以上这些合成生物学的优化策略，大多数WCMB都可以实现重金属的痕量检测。

3 总结与展望

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基于不同类型转录因子的WCMB在环境重金属污染检测方面展现出极为广阔的应用潜力。由于重金属如Pb、Hg、Cd、As等对人体健康和生态环境系统构成严重威胁，因此对环境中重金属进行准确、快速、高效且灵敏的检测对于保护环境和人体健康具有重要的意义。这种检测技术不仅能确定重金属污染的种类与浓度，还为重金属污染的防控提供有力支持。此外，借助合成生物学的策略，可以对WCMB的结构和功能进行改造和优化，从而实现对低浓度重金属污染物的精准检测，对于及时发现和解决潜在的重金属环境污染问题具有重要意义。此外，WCMB还具有实时在线监测的潜力，通过将WCMB集成到常规环境监测系统中，可以实现对重金属污染物的实时、连续监测，为环境保护部门提供及时、准确的数据支持，有助于制定科学的污染防控策略。

然而，与一些化学检测仪器(如ICP-MS等)相比，尽管化学仪器成本较高、操作相对复杂，但其准确性高；相比之下，WCMB在成本上更具优势，但在准确性方面仍有待提升。其次，由于现有合成生物学技术的局限性和转基因生物可能带来的安全威胁等问题，目前WCMB的设计过程和实际应用仍存在困难。不同类型转录因子的选择和应用仍需更多实验验证和优化。此外，WCMB的稳定性和可重复性问题也亟待解决，将其从实验室规模转移到商业规模同样面临重大挑战。未来可以继续提高基于转录因子的WCMB的灵敏度和特异性，降低WCMB的制造成本和操作复杂性，并优化其稳定性和再生性，以实现实时连续监测。此外，WCMB的设计可以更加注重创新和智能化。一方面，通过开发新的识别元件，有望为重金属污染的有效防控提供更有力的技术支持；另一方面，随着互联网、人工智能等技术的不断发展，重金属WCMB可以向小巧、便携、灵敏、可重复使用等方向进一步发展。随着相关技术和法规的不断进步与完善，相信这一领域将会取得更加丰硕的成果，为保护环境和人体健康提供更加有力的保障。

基金

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广东省重点领域研发计划(2023B0202040001)
广东省科学院发展专项资金(2022GDASZH-2022010101)

参考文献

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文献

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参考文献引证文献

排序方式：

[1]

VELUSAMY

, PERIYASAMY

, KUMAR

, RANGASAMY

, PAULINE

JMN

, RAMARAJU

, MOHANASUNDARAM

, VO DV. Biosensor for heavy metals detection in wastewater: a review[J]. Food and Chemical Toxicology, 2022, 168: 113307.

[2]

KARTHIK

, SELVAKUMAR

, KUMAR

, SATHEESKUMAR

, VIJAYSUNDER

, HARIHARAN

, ANTONY

. Recent advances in electrochemical sensor developments for detecting emerging pollutant in water environment[J]. Chemosphere, 2022, 304: 135331.

[3]

张祖烨, 何梦园, 乔月, 贾润梅, 彭祥轩, 麦华明, 陈思. Agromyces sp. CS16去除水体中重金属镉、镍、铜、锌离子的研究[J]. 微生物学报, 2023, 63(7): 2791-2808.

ZHANG

, HE

, QIAO

, JIA

, PENG

, MAI HM, CHEN

. Removal of cadmium, nickel, copper, and zinc ions from water by Agromyces sp. CS16[J]. Acta Microbiologica Sinica, 2023, 63(7): 2791-2808 (in Chinese).

[4]

BRIFFA

, SINAGRA

, BLUNDELL

. Heavy metal pollution in the environment and their toxicological effects on humans[J]. Heliyon, 2020, 6(9): e04691.

[5]

ALI Z, YOUSAFZAI

, SHER

, MUHAMMAD

, NAYAB

, AQEEL

SAM

, SHAH

, ASCHNER

, KHAN

. Toxicity and bioaccumulation of manganese and chromium in different organs of common carp (Cyprinus carpio) fish[J]. Toxicology Reports, 2021, 8: 343-348.

[6]

, LU

, LI

, ZENG

, SHEN

, YU

, LI

. Microbial extracellular polymeric substances alleviate cadmium toxicity in rice (Oryza sativa L.) by regulating cadmium uptake, subcellular distribution and triggering the expression of stress-related genes[J]. Ecotoxicology and Environmental Safety, 2023, 257: 114958.

[7]

ZHANG

, YANG

, LAN

, HUANG

, ZHANG

, HUANG

, YANG

, RU

. Water quality degradation due to heavy metal contamination: health impacts and eco-friendly approaches for heavy metal remediation[J]. Toxics, 2023, 11(10): 828.

[8]

KOMIJANI

, SHAMABADI

, SHAHIN

, EGHBALPOUR

, TAHSILI

, BAHRAM

. Heavy metal pollution promotes antibiotic resistance potential in the aquatic environment[J]. Environmental Pollution, 2021, 274: 116569.

[9]

NAZ S, FAZIO

, HABIB

, NAWAZ

, ATTAULLAH

, ULLAH

, HAYAT

, AHMED

. Incidence of heavy metals in the application of fertilizers to crops (wheat and rice), a fish (common carp) pond and a human health risk assessment[J]. Sustainability, 2022, 14(20): 13441.

[10]

UNITED

STATES ENVIRONMENTAL PROTECTION AGENCY

. Stronger protections from lead in drinking water: next steps for the lead and copper rule [EB/OL]. (2021-12-01)[2024-3-20]. https://www.epa.gov/system/files/documents/2021-12/lcrr-review-fact-sheet_0.pdf

[11]

EUROPEAN

COMMISSION

. Water framework directive[EB/OL]. (2023-12-20)[2024-3-20]. https://environment.ec.europa.eu/topics/water/water-framework-directive_en

[12]

ARJOMANDI

, SHIRKHANLOO

. A review: analytical methods for heavy metals determination in environment and human samples[J]. Analytical Methods in Environmental Chemistry Journal, 2019, 2(3): 97-126.

[13]

MATHUR

, SINGH

, RANJAN

. Genetic circuits in microbial biosensors for heavy metal detection in soil and water[J]. Biochemical and Biophysical Research Communications, 2023, 652: 131-137.

[14]

KIM

, RASTON

NHA

, GU

. Aptamer-based nanobiosensors[J]. Biosensors and Bioelectronics, 2016, 76: 2-19.

[15]

REBOLLAR-PÉREZ

, CAMPOS-TERÁN

, ORNELAS-SOTO

, MÉNDEZ-ALBORES

, TORRES

. Biosensors based on oxidative enzymes for detection of environmental pollutants[J]. Biocatalysis, 2016, 1(1): 118-129.

[16]

HOSSAIN

, SLAUGHTER

. Flexible electrochemical uric acid and glucose biosensor[J]. Bioelectrochemistry, 2021, 141: 107870.

[17]

, JIANG

, XUE

, PENG

, SHI

, ZHANG

, LI

, PAN

, WANG

, FENG

, QIAO

, CHEN

, LUO

, CHEN

. Recent progress in biosensors for detection of tumor biomarkers[J]. Molecules, 2022, 27(21): 7327.

[18]

GAVRILAȘ

, CȘ

URSACHI

, PERȚA-CRIȘAN

, MUNTEANU

. Recent trends in biosensors for environmental quality monitoring[J]. Sensors, 2022, 22(4): 1513.

[19]

MAJER-BARANYI

, SZÉKÁCS

, ADÁNYI

. Application of electrochemical biosensors for determination of food spoilage[J]. Biosensors, 2023, 13(4): 456.

[20]

MORASKIE

, ROSHID

MHO

, O’CONNOR

, DIKICI

, ZINGG

, DEO

, DAUNERT

. Microbial whole-cell biosensors: current applications, challenges, and future perspectives[J]. Biosensors and Bioelectronics, 2021, 191: 113359.

[21]

BEREZA-MALCOLM

, MANN

, FRANKS

. Environmental sensing of heavy metals through whole cell microbial biosensors: a synthetic biology approach[J]. ACS Synthetic Biology, 2015, 4(5): 535-546.

[22]

LIU

, YU

, ZHANG

, LIU

, LI

, SONG

. Engineering whole-cell microbial biosensors: design principles and applications in monitoring and treatment of heavy metals and organic pollutants[J]. Biotechnology Advances, 2022, 60: 108019.

[23]

DING

, ZHOU

, DENG

. Transcription-factor-based biosensor engineering for applications in synthetic biology[J]. ACS Synthetic Biology, 2021, 10(5): 911-922.

[24]

HOSSAIN

, SAINI

, MIYAKE

, LING

, CHANG

. Genetic biosensor design for natural product biosynthesis in microorganisms[J]. Trends in Biotechnology, 2020, 38(7): 797-810.

[25]

, XU

, JIN

, LIU

, LI

, DU

, LV

, LIU

. Genetically encoded biosensors for microbial synthetic biology: from conceptual frameworks to practical applications[J]. Biotechnology Advances, 2023, 62: 108077.

[26]

, JACOBSEN

, GIEDROC

. Coordination chemistry of bacterial metal transport and sensing[J]. Chemical reviews, 2009, 109(10): 4644-4681.

[27]

TULIN

, FIGUEROA

, CHECA

, SONCINI

. The multifarious MerR family of transcriptional regulators[J]. Molecular Microbiology, 2024, 121(2): 230-242.

[28]

DOS REIS FERREIRA

, PIRES

, RIBEIRO

, CARLIER

, COSTA

, SCHWAN

, SILVA

. Impact of lead (Pb²⁺) on the growth and biological activity of Serratia marcescens selected for wastewater treatment and identification of its zntR gene: a metal efflux regulator[J]. World Journal of Microbiology and Biotechnology, 2023, 39(4): 91.

[29]

EKAS

, WANG

, SILVERMAN

, LUCKS

, KARIM

, JEWETT

. Engineering a PbrR-based biosensor for cell-free detection of lead at the legal limit[J]. ACS Synthetic Biology, 2024, 13(9): 3003-3012.

[30]

GUO

, CHEN

, SU

, CHEN

, LIU

, SUN

. High-throughput visualization mutation screening technology to enhance the specificity of CadR based whole-cell cadmium biosensor[J]. Biosensors and Bioelectronics, 2024, 256: 116266.

[31]

KANDARI

, JOSHI

. PerR: a peroxide sensor eliciting metal ion-dependent regulation in various bacteria[J]. Molecular Biotechnology, 2024: 1-12.

[32]

SEVILLA

, BES MT, PELEATO

, FILLAT

. Fur-like proteins: beyond the ferric uptake regulator (Fur) paralog[J]. Archives of Biochemistry and Biophysics, 2021, 701: 108770.

[33]

BARROWS

, STUBBS

, PADILLA-MONTOYA

, LEEPER

, DYKE

. Manganese-dependent transcription regulation by MntR and PerR in Thermus thermophilus HB8[J]. Molecular Microbiology, 2024, 121(6): 1228-1244.

[34]

MARCOS-TORRES

, JUNIAR

, GRIESE

. The molecular mechanisms of the bacterial iron sensor IdeR[J]. Biochemical Society Transactions, 2023, 51(3): 1319-1329.

[35]

JELIĆ MATOŠEVIĆ

, RADMAN

, LOUBSER

, CRNOLATAC

, PIANTANIDA

, CUKROWSKI

, LEŠČIĆ AŠLER

, BERTOŠA

. Structural dynamics of the Bacillus subtilis MntR transcription factor is locked by Mn²⁺ binding[J]. International Journal of Molecular Sciences, 2023, 24(2): 957.

[36]

ZHENG

, WEI

, QIU

, JIA

, ZHOU

, JIAO

. TroR negatively regulates the TroABCD system and is required for resistance to metal toxicity and virulence in Streptococcus suis [J]. Applied and Environmental Microbiology, 2021, 87(20): e01375-21.

[37]

CAO

, LI

, WANG

, HU

, XIANG

, ZHANG

, LIU

. Cellular uptake of nickel by NikR is regulated by phase separation[J]. Cell Reports, 2023, 42(6): 112518.

[38]

ZAMBELLI

. Intracellular phase separation and its role in nickel sensing[J]. Trends in Cell Biology, 2023, 33(9): 732-733.

[39]

RIGOLI

, POTESTIO

, MENICHETTI

. A multi-scale analysis of the CzrA transcription repressor highlights the allosteric changes induced by metal ion binding[EB/OL]. 2024. https://arxiv.org/abs/2409.03584v1

[40]

GALLO

, ANTONUCCI

, PIRONE

, AMORESANO

, CONTURSI

, LIMAURO

, PEDONE

, BARTOLUCCI

, FIORENTINO

. A physicochemical investigation on the metal binding properties of TtSmtB, a thermophilic member of the ArsR/SmtB transcription factor family[J]. International Journal of Biological Macromolecules, 2019, 138: 1056-1063.

[41]

QIU

, TONG

, YANG

, FENG

. Molecular insights into a novel Cu(I)-sensitive ArsR/SmtB family repressor in extremophile Acidithiobacillus caldus [J]. Applied and Environmental Microbiology, 2023, 89(1): e01266-22.

[42]

CHEN

, ZHANG

, LI

, WANG

, YE

. De novo design of the ArsR regulated P _ars promoter enables a highly sensitive whole-cell biosensor for arsenic contamination[J]. Analytical Chemistry, 2022, 94(20): 7210-7218.

[43]

KUMARI

WMNH

, THIRUCHITTAMPALAM

, WEERASINGHE

MSS

, CHANDRASEKHARAN

, WIJAYARATHNA

. Characterization of a Bacillus megaterium strain with metal bioremediation potential and in silico discovery of novel cadmium binding motifs in the regulator, CadC[J]. Applied Microbiology and Biotechnology, 2021, 105(6): 2573-2586.

[44]

, CHEN

, LIAO

, HUI

, LI

, HE

. A novel stress-inducible CmtR-ESX3-Zn²⁺ regulatory pathway essential for survival of Mycobacterium bovis under oxidative stress[J]. Journal of Biological Chemistry, 2020, 295(50): 17083-17099.

[45]

IGBARIA-JABER

, HOFMANN

, GEVORKYAN-AIRAPETOV

, SHENBERGER

, RUTHSTEIN

. Revealing the DNA binding modes of CsoR by EPR spectroscopy[J]. ACS Omega, 2023, 8(42): 39886-39895.

[46]

, VAVRA

, CARR

, HUANG

, MARONEY

, WILMOT

. Complexation of the nickel and cobalt transcriptional regulator RcnR with DNA[J]. Acta Crystallographica Section F: Structural Biology Communications, 2020, 76(1): 25-30.

[47]

HAN

, CUI

, LIN

, CHEN

, SUO

, MA

, WANG

, HAO

, CHENG

, ZHOU

. Efficient overproduction of active nitrile hydratase by coupling expression induction and enzyme maturation via programming a controllable cobalt-responsive gene circuit[J]. Frontiers in Bioengineering and Biotechnology, 2020, 8: 193.

[48]

HIRSCH

, HOFMANN

, SHENBERGER

, GEVORKYAN-AIRAPETOV

, RUTHSTEIN

. Conformations and local dynamics of the CopY metal sensor revealed by EPR spectroscopy[J]. Biochemistry, 2023, 62(3): 797-807.

[49]

NAM KH. Fluorescent protein-based metal biosensors[J]. Chemosensors, 2023, 11(4): 216.

[50]

BOGDANOVA

, ZAITSEVA

, SMIRNOV

, BALEEVA

, GAVRIKOV

, MYASNYANKO

, GONCHARUK

, KOT EF, MINEEV

, MISHIN

, BARANOV

. NanoLuc luciferase as a fluorogen-activating protein for GFP chromophore based fluorogens[J]. International Journal of Molecular Sciences, 2023, 24(9): 7958.

[51]

, DONG

, QU

, HAN

, YOU

, LI

, HAN

. An enzyme-fused phycobiliprotein synthesis system developed for visual whole-cell biosensors for the detection of cadmium during wastewater treatment[J]. Environmental Technology & Innovation, 2024, 36: 103882.

[52]

SINGH

, KUMAR

. Recent advances in synthetic biology-enabled and natural whole-cell optical biosensing of heavy metals[J]. Analytical and Bioanalytical Chemistry, 2021, 413(1): 73-82.

[53]

, MELIANA

, MUNAWAROH

HSH

, KARAMAN

, KARIMI-MALEH

, LOW SS, SHOW

. Recent advances in the analytical strategies of microbial biosensor for detection of pollutants[J]. Chemosphere, 2022, 306: 135515.

[54]

ZHU

, XIANG

, CHEN

, WANG

, JIANG

, HAO

, ZHANG

, WANG

, LI

, OMER

, ZHANG

, WANG

, ZHUANG

, HUANG

. Engineered Bacillus subtilis Biofilm@ Biochar living materials for in-situ sensing and bioremediation of heavy metal ions pollution[J]. Journal of Hazardous Materials, 2024, 465: 133119.

[55]

FAN

, ZHANG

, MO

, YUAN

. Engineering Saccharomyces cerevisiae-based biosensors for copper detection[J]. Microbial Biotechnology, 2022, 15(11): 2854-2860.

[56]

LIU

, TANG

, HU

. Application of cyanobacteria as chassis cells in synthetic biology[J]. Microorganism, 2024, 12(7): 1375.

[57]

WANG

, CHENG

, YANG

, WEI

, XIA

, WANG

, BEN Y, ZHOU

, YANG

, HUANG

. A living plant cell-based biosensor for real-time monitoring invisible damage of plant cells under heavy metal stress[J]. Science of the Total Environment, 2019, 697: 134097.

[58]

JEON

, LEE

, JANG

, KIM

, YOON

. Design of Pb(II)-specific E. coli-based biosensors by engineering regulatory proteins and host cells[J]. Frontiers in Microbiology, 2022, 13: 881050.

[59]

KIM

, KIM

, SUBHADRA

, WOO SG, RHA E, KIM

, KIM

, LEE

. A genetically encoded biosensor for monitoring isoprene production in engineered Escherichia coli [J]. ACS Synthetic Biology, 2018, 7(10): 2379-2390.

[60]

GUO

, WANG

, DU

, LIU

, CHI

, HE

, HUANG

, LUO

, XU

. A test strip platform based on a whole-cell microbial biosensor for simultaneous on-site detection of total inorganic mercury pollutants in cosmetics without the need for predigestion[J]. Biosensors and Bioelectronics, 2020, 150: 111899.

[61]

ZEVALLOS-ALIAGA

, de GRAEVE

, OBANDO-CHÁVEZ

, VACCARI

, GAO

, PEETERS

, GUERRA

. Highly sensitive whole-cell mercury biosensors for environmental monitoring[J]. Biosensors, 2024, 14(5): 246.

[62]

KHAN

, SALAMA

, CHEN

, NI

HYH

, ZHAO

, ZHOU

, PEI

, SANI

, LING

, LIU

, LI

. A novel biosensor for zinc detection based on microbial fuel cell system[J]. Biosensors and Bioelectronics, 2020, 147: 111763.

[63]

HUI

, GUO

, LIU

, ZHANG

, GAO

, YANG

, YI

. Genetic control of violacein biosynthesis to enable a pigment-based whole-cell lead biosensor[J]. RSC Advances, 2020, 10(47): 28106-28113.

[64]

NOURMOHAMMADI

, HOSSEINKHANI

, NEDAEINIA

, KHOSHDEL-SARKARIZI

, NEDAEINIA

, RANJBAR

, EBRAHIMI

, FARJAMI

, NOURMOHAMMADI

, MAHMOUDI

, GOLI

, FERNS

, SADEGHIZADEH

. Construction of a sensitive and specific lead biosensor using a genetically engineered bacterial system with a luciferase gene reporter controlled by pbr and cadA promoters[J]. Biomedical Engineering Online, 2020, 19(1): 79.

[65]

HUI

, GUO

, LI

, GAO

, YI

. Detection of environmental pollutant cadmium in water using a visual bacterial biosensor[J]. Scientific Reports, 2022, 12(1): 6898.

[66]

BEREZA-MALCOLM

, ARACIC

, KANNAN

, MANN

, FRANKS

. Functional characterization of Gram-negative bacteria from different genera as multiplex cadmium biosensors[J]. Biosensors and Bioelectronics, 2017, 94: 380-387.

[67]

GUO

, CHEN

, LIN

, CHEN

, LEE

, YEH

. Determination of gold ions in human urine using genetically engineered microorganisms on a paper device[J]. ACS Sensors, 2018, 3(4): 744-748.

[68]

FANG

, ZHU

, CHEN

, WANG

, XU

, SUN

, GUO

, YOO

, TIE CJ, JIANG

, LI

. Copy number of ArsR reporter plasmid determines its arsenite response and metal specificity[J]. Applied Microbiology and Biotechnology, 2018, 102(13): 5753-5761.

[69]

, WANG

, YUAN

, LIU

, FU

, SUN

, DANG

, HOLMES

. Development of a whole-cell biosensor based on an ArsR-Pars regulatory circuit from Geobacter sulfurreducens [J]. Environmental Science and Ecotechnology, 2021, 6: 100092.

[70]

CHEN

, YAO

, SONG

, LIN

, ZHOU

, YUAN

, SONG

, SUN

, XU

. A novel antimony-selective ArsR transcriptional repressor and its specific detection of antimony trioxide in environmental samples via bacterial biosensor[J]. Biosensors and Bioelectronics, 2023, 220: 114838.

[71]

HUI

, GUO

, LI

, CHEN

, YI

. Differential detection of bioavailable mercury and cadmium based on a robust dual-sensing bacterial biosensor[J]. Frontiers in Microbiology, 2022, 13: 846524.

[72]

HUI

, GUO

, WU

, LIU

, YANG

, GUO

, XIE

, YI

. Detection of bioavailable cadmium by double-color fluorescence based on a dual-sensing bioreporter system[J]. Frontiers in Microbiology, 2021, 12: 696195.

[73]

HUI

, GUO

, LI

, LIU

, CHEN

, YI

, ZHANG

. Indigoidine biosynthesis triggered by the heavy metal-responsive transcription regulator: a visual whole-cell biosensor[J]. Applied Microbiology and Biotechnology, 2021, 105(14/15): 6087-6102.

[74]

, HUI

, WU

, GAO

, HUANG

, GUO

. Dual-colored bacterial biosensor responsive to cadmium, mercury, and lead for detecting heavy metal pollution in seawater[J]. Ecological Indicators, 2024, 166: 112244.

[75]

PARK

, OVERTON

, LIOU

, JIAO

. Identification of a U/Zn/Cu responsive global regulatory two‐component system in Caulobacter crescentus [J]. Molecular Microbiology, 2017, 104(1): 46-64.

[76]

PARK

, TAFFET

. Combinatorial sensor design in Caulobacter crescentus for selective environmental uranium detection[J]. ACS Synthetic Biology, 2019, 8(4): 807-817.

[77]

FANG

, ZHANG

. Bacterial MerR family transcription regulators: activationby distortion: the mechanism of transcription regulation by MerR[J]. Acta Biochimica et Biophysica Sinica, 2021, 54(1): 25.

[78]

, WEI

, YU

, DENG

, XU

, MA

, ZHANG

. Expression of cadR enhances its specific activity for Cd detoxification and accumulation in Arabidopsis[J]. Plant and Cell Physiology, 2016, 57(8): 1720-1731.

[79]

LEE

, KIM

, JANG

, KIM

, YOON

. Antimony sensing whole-cell bioreporters derived from ArsR genetic engineering[J]. Applied Microbiology and Biotechnology, 2020, 104: 2691-2699.

[80]

CHEN

, ROSEN

. Biosensors for inorganic and organic arsenicals[J]. Biosensors, 2014, 4(4): 494-512.

[81]

SALTIKOV

, OLSON

. Homology of Escherichia coli R773 arsA, arsB, and arsC genes in arsenic-resistant bacteria isolated from raw sewage and arsenic-enriched creek waters[J]. Applied and Environmental Microbiology, 2002, 68(1): 280-288.

[82]

SANTHA

, PANDARANAYAKA

EPJ

, ROSEN

, THIYAGARAJAN

. Purification, crystallization and preliminary X-ray diffraction studies of the arsenic repressor ArsR from Corynebacterium glutamicum [J]. Acta Crystallographica Section F: Structural Biology and Crystallization Communications, 2011, 67(12): 1616-1618.

[83]

QIN

, FU

, YE

, BENCZE

, STEMMLER

, RAWLINGS

, ROSEN

. Convergent evolution of a new arsenic binding site in the ArsR/SmtB family of metalloregulators[J]. Journal of Biological Chemistry, 2007, 282(47): 34346-34355.

[84]

CHEN

, NADAR

, ROSEN

. A novel MAs (III)‐selective ArsR transcriptional repressor[J]. Molecular Microbiology, 2017, 106(3): 469-478.

[85]

KHAN

, SHEN

, FATMI

, CAMPBELL

, BOKHARI

. Design and prototyping of genetically encoded arsenic biosensors based on transcriptional regulator AfArsR[J]. Biomolecules, 2021, 11(9): 1276.

[86]

MUSIANI

, ZAMBELLI

, BAZZANI

, MAZZEI

, CIURLI

. Nickel-responsive transcriptional regulators[J]. Metallomics, 2015, 7(9): 1305-1318.

[87]

YOON

, KANG

, CHAE

, KIM

, LEE

, JEONG

, AN

. Arsenic bioavailability in soils before and after soil washing: the use of Escherichia coli whole-cell bioreporters[J]. Environmental Science and Pollution Research, 2016, 23(3): 2353-2361.

[88]

GONZALEZ-FLO

, ALABALL

, MACIA

. Two-component biosensors: unveiling the mechanisms of predictable tunability[J]. ACS Synthetic Biology, 2020, 9(6): 1328-1335.

[89]

QIN

, LIU

, XU

, LI

. Mining and design of biosensors for engineering microbial cell factory[J]. Current Opinion in Biotechnology, 2022, 75: 102694.

[90]

KIM

, JANG

, KIM

, YOON

. Modulation of the metal (loid) specificity of whole-cell bioreporters by genetic engineering of ZntR metal-binding loops[J]. Journal of Microbiology and Biotechnology, 2020, 30(5): 681-688.

[91]

LEE

, JEON

, JANG

, YOON

. Derivation of Pb(II)-sensing Escherichia coli cell-based biosensors from arsenic responsive genetic systems[J]. AMB Express, 2021, 11: 169.

[92]

MENDOZA

, SONCINI

, CHECA

. Engineering of a Au-sensor to develop a Hg-specific, sensitive and robust whole-cell biosensor for on-site water monitoring[J]. Chemical Communications, 2020, 56(48): 6590-6593.

[93]

GHATAORA

, GEBHARD

, REEKSTING

. Chimeric MerR-family regulators and logic elements for the design of metal sensitive genetic circuits in Bacillus subtilis [J]. ACS Synthetic Biology, 2023, 12(3): 735-749.

[94]

CAI

, ZHU

, SHEN

, MA

, BAO

, CHEN

, WEI

, LIU

, WU

, CHEN

. Evolved biosensor with high sensitivity and specificity for measuring cadmium in actual environmental samples[J]. Environmental Science & Technology, 2022, 56(14): 10062-10071.

[95]

GUO

, DU

, XIE

, HE

, HUANG

, LUO

, XU

. Using the promoters of MerR family proteins as “rheostats” to engineer whole-cell heavy metal biosensors with adjustable sensitivity[J]. Journal of Biological Engineering, 2019, 13(1): 70.

[96]

ZHU

, CHEN

, CAI

, ZHANG

, MA

, BAO

, ZHAO

, WU

, CHEN

. Engineering the ultrasensitive visual whole-cell biosensors by evolved MerR and 5′ UTR for detection of ultratrace mercury[J]. Environmental Science & Technology, 2023, 57(44): 16964-16973.

[97]

CHEN

, WEI

, YIN

, TONG

, LU

, YE

. Development of a highly sensitive whole-cell biosensor for arsenite detection through engineered promoter modifications[J]. ACS Synthetic Biology, 2019, 8(10): 2295-2302.

[98]

CHEN

, JIANG

, TIE CJ, YOO

, WANG

, XU

, SUN

, GUO

, LI

. Contribution of nonconsensus base pairs within ArsR binding sequences toward ArsR-DNA binding and arsenic-mediated transcriptional induction[J]. Journal of Biological Engineering, 2019, 13: 53.

[99]

, JIANG

, XU

, FANG

, WANG

, SUN

, GUO

. Identification of stress-responsive transcription factors with protein-bound Escherichia coli genomic DNA libraries[J]. AMB Express, 2020, 10: 199.

[100]

JIA

, BU

, ZHAO

, WU

. Sensitive and specific whole-cell biosensor for arsenic detection[J]. Applied and Environmental Microbiology, 2019, 85(11): e00694-19.

[101]

WAN

, VOLPETTI

, PETROVA

, FRENCH

, MAERKL

, WANG

. Cascaded amplifying circuits enable ultrasensitive cellular sensors for toxic metals[J]. Nature Chemical Biology, 2019, 15(5): 540-548.

[102]

JIA

, LIU

, MA

, WU

. Construction of cadmium whole-cell biosensors and circuit amplification[J]. Applied Microbiology and Biotechnology, 2021, 105(13): 5689-5699.

[103]

ZHANG

, HU

, JIA

. Highly sensitive whole-cell biosensor for cadmium detection based on a negative feedback circuit[J]. Frontiers in Bioengineering and Biotechnology, 2021, 9: 799781.

[104]

WANG

, ZHANG

, TAO

, LV

, DENG

, LI

. E. coli biosensor based on modular GFP and LuxI/LuxR cyclic amplification circuit for sensitive detection of lysine[J]. Analytical and Bioanalytical Chemistry, 2022, 414(29): 8299-8307.

2025年第65卷第3期

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出版时间：2025-03-04

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出版历史

收稿日期：2024-10-10
录用日期：2024-11-25

基金

Key Research and Development Program of Guangdong Province(2023B0202040001)

广东省重点领域研发计划(2023B0202040001)

GDAS Project of Science and Technology Development(2022GDASZH-2022010101)

广东省科学院发展专项资金(2022GDASZH-2022010101)

作者信息

1 河南科技大学食品与生物工程学院，河南洛阳

2 广东省科学院微生物研究所，华南应用微生物国家重点实验室，广东省菌种保藏与应用重点实验室，广东广州

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https://castjournals.cast.org.cn/joweb/wswxb/CN/10.13343/j.cnki.wsxb.20240619

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2种不同金属材料的力学参数

科 Family	属数 Number of genus	种数 Number of species	占总种数比例 Percentage of total species (%)	属 Genus	种数 Number of species	占总种数比例 Percentage of total species (%)
鹅膏菌科Amanitaceae	2	11	5.26	鹅膏菌属 Amanita	10	4.78
小菇科 Mycenaceae	2	12	5.74	丝盖伞属 Inocybe	5	2.39
多孔菌科 Polyporaceae	8	14	6.70	蜡蘑属 Laccaria	5	2.39
红菇科 Russulaceae	3	23	11.00	小皮伞属 Marasmius	6	2.87
				小菇属 Mycena	11	5.26
				光柄菇属 Pluteus	5	2.39
				红菇属 Russula	17	8.13
				栓菌属 Trametes	5	2.39

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Metal-responsive protein family	Family members and responders	References
MerR family	MerR (Hg), ZntR (Cd, Pb, Zn), ZccR (Zn, PbrR (Pb), CueR (Cu, Au), GolS (Au), HmrR (Ag), CoaR (Co), CadR (Cd), NimR (Ni)	[27-30]
Fur family	Fur (Fe), Zur (Zn), Mur (Mn), Nur (Ni), PerR (Mn/Fe)	[31-32]
DtxR family	DtxR, IdeR, SirR (Fe), MntR (Mn, ScaR (Cd), TroR (Mn, Zn)	[33-36]
NikR family	NikR (Ni)	[37-38]
ArsR/SmtB family	CmtR, ZiaR, AztR (Zn), CzrA (Zn, Co), ArsR (As, Sb, Bi), CadC (Cd, Pb, Zn), SmtB (As, Cd), NmtR (Ni, Co), KmtR (Ni), BxmR (Cu, Ag, Zn, Cd)	[39-44]
CsoR-RcnR family	CsoR (Cu), RcnR (Ni、Co)	[45-47]
CopY family	CopY (Cu)	[48]

Metal-responsive protein family

Family members and responders

References

MerR family

MerR (Hg), ZntR (Cd, Pb, Zn), ZccR (Zn, PbrR (Pb), CueR (Cu, Au), GolS (Au), HmrR (Ag), CoaR (Co), CadR (Cd), NimR (Ni)

[27-30]

Fur family

Fur (Fe), Zur (Zn), Mur (Mn), Nur (Ni), PerR (Mn/Fe)

[31-32]

DtxR family

DtxR, IdeR, SirR (Fe), MntR (Mn, ScaR (Cd), TroR (Mn, Zn)

[33-36]

NikR family

NikR (Ni)

[37-38]

ArsR/SmtB family

CmtR, ZiaR, AztR (Zn), CzrA (Zn, Co), ArsR (As, Sb, Bi), CadC (Cd, Pb, Zn), SmtB (As, Cd), NmtR (Ni, Co), KmtR (Ni), BxmR (Cu, Ag, Zn, Cd)

[39-44]

CsoR-RcnR family

CsoR (Cu), RcnR (Ni、Co)

[45-47]

CopY family

CopY (Cu)

[48]

Sensing element	Detection object	Reporter gene	Host cell	Application	Detection range (μmol/L)	References
Transcription activator
MerR	Hg²⁺	rfp	Escherichia coli DH5α	Rapid and convenient screening of total inorganic mercury in cosmetics	0.050 0-10	[60]
	Hg²⁺	rfp, amilcpblue	Escherichia coli DH5α	The extremely wide linear range meets the different monitoring requirements	0.001 0-1 (Mer-RFP), 0.002 0-0.125 0 (Mer-Blue)	[61]
ZntR	Zn²⁺	zntR, ribB, oprF	Escherichia coli BL21	Microbial fuel cell electrobiosensor	0-400	[62]
PbrR	Pb²⁺	vio ABCDE	Escherichia coli TOP10	Detection of biologically available Pb²⁺ in water	0.187 5-1.500 0	[63]
	Pb²⁺	Luc	Escherichia coli DH5α	Detection of Pb²⁺ in environmental and biological samples	1-100	[64]
CadR	Cd²⁺	vio ABCDE	Escherichia coli TOP10	Detection of soluble Cd²⁺ in environmental water samples	0.049 0-25	[65]
	Cd²⁺	rfp	Multiple Gram-negative bacteria	The importance of WCMB testing in different genera of bacteria was emphasized	0.500 0–2.000 0 μg/mL (E. coli DH5α) 0.100 0 μg/mL (Pseudomonas aeruginosa PAO1) 10 μg/mL (Shewanella oneidensis MR-1) 0.250 0 μg/mL (Enterobacter sp. NCR3) 1 μg/mL (Enterobacter sp. LCR17)	[66]
	Pb²⁺	Luc	Escherichia coli DH5α	For the detection of Pb²⁺ in environmental and biological samples	0.010 0-10	[64]
CueR	Au³⁺	rfp	Cupriavidus etallidurans	Monitoring the concentration of Au³⁺ in human urine	≥0.110 0	[67]
Transcription inhibitors
EcArsR	AsO₃^3-	luc	Escherichia coli DH5α	A WCMB that is highly sensitive to arsenite has been developed	0.100 0-1≥0.040 0	[68]
ArsR1	AsO₃^3-	gfp	Escherichia coli TOP10	A WCMB that is highly sensitive to arsenite has been developed	0.030 0-0.100 0 (2.250 0-7.500 0 μg/L) ≥0.010 0	[69]
SxArsR	Sb³⁺	luc	Sphingobium xenophagum C1	A novel subtype of the ArsR family transcription factor, designated as SxArsR, has been identified, which exhibits specific responsiveness to Sb	0.010 0-6 ≥0.010 0^*	[70]
Combination of transcription factors
MerR, CadR	Hg²⁺, Cd²⁺	egfp, mCherry	Escherichia coli TOP10	Detection of concurrent heavy metal contaminants in the environment	0-40 (Hg²⁺), 0-200 (Cd²⁺)	[71]
CadC, CadR	Cd²⁺	egfp and mCherry	Escherichia coli TOP10	Detection of high concentrations of biologically available Cd in water	≥0.050 0 (CadC-eGFP), ≥0.100 0 (CadR-mCherry)	[72]
MerR, PbrR	Hg²⁺, Pb²⁺	Indigoidine biosynthetic module	Escherichia coli TOP10	Detection of Hg²⁺ and Pb²⁺ in environmental samples	≥0.008 0	[73]
CadR, MerR	Cd²⁺, Pb²⁺, Hg²⁺	vioABE, vioC	Escherichia coli TOP10	Designed for detecting heavy metal contaminants in seawater	0.004 9-40, ≥0.004 9 (Cd²⁺), 0.024 4-200, ≥0.024 4 (Pb²⁺), 0.003 7-0.468 8, ≥0.000 5 (Hg²⁺)^*	[74]
UzcRS, UrpRS	UO₂²⁺	gfp	Caulobacter vibrioides	Detection of uranium (U) in groundwater samples	≥1	[75-76]

Sensing element

Detection object

Reporter gene

Host cell

Application

Detection range

(μmol/L)

References

Transcription activator

MerR

Hg²⁺

rfp

Escherichia coli DH5α

Rapid and convenient screening of total inorganic mercury in cosmetics

0.050 0-10

[60]

Hg²⁺

rfp, amilcpblue

Escherichia coli DH5α

The extremely wide linear range meets the different monitoring requirements

0.001 0-1 (Mer-RFP), 0.002 0-0.125 0 (Mer-Blue)

[61]

ZntR

Zn²⁺

zntR, ribB, oprF

Escherichia coli BL21

Microbial fuel cell electrobiosensor

0-400

[62]

PbrR

Pb²⁺

vio ABCDE

Escherichia coli TOP10

Detection of biologically available Pb²⁺ in water

0.187 5-1.500 0

[63]

Pb²⁺

Luc

Escherichia coli DH5α

Detection of Pb²⁺ in environmental and biological samples

1-100

[64]

CadR

Cd²⁺

vio ABCDE

Escherichia coli TOP10

Detection of soluble Cd²⁺ in environmental water samples

0.049 0-25

[65]

Cd²⁺

rfp

Multiple Gram-negative bacteria

The importance of WCMB testing in different genera of bacteria was emphasized

0.500 0–2.000 0 μg/mL (E. coli DH5α)

0.100 0 μg/mL (Pseudomonas aeruginosa PAO1)

10 μg/mL (Shewanella oneidensis MR-1)

0.250 0 μg/mL (Enterobacter sp. NCR3)

1 μg/mL (Enterobacter sp. LCR17)

[66]

Pb²⁺

Luc

Escherichia coli DH5α

For the detection of Pb²⁺ in environmental and biological samples

0.010 0-10

[64]

CueR

Au³⁺

rfp

Cupriavidus etallidurans

Monitoring the concentration of Au³⁺ in human urine

≥0.110 0

[67]

Transcription inhibitors

EcArsR

AsO₃^3-

luc

Escherichia coli DH5α

A WCMB that is highly sensitive to arsenite has been developed

0.100 0-1≥0.040 0

[68]

ArsR1

AsO₃^3-

gfp

Escherichia coli TOP10

A WCMB that is highly sensitive to arsenite has been developed

0.030 0-0.100 0

(2.250 0-7.500 0 μg/L)

≥0.010 0

[69]

SxArsR

Sb³⁺

luc

Sphingobium xenophagum C1

A novel subtype of the ArsR family transcription factor, designated as SxArsR, has been identified, which exhibits specific responsiveness to Sb

0.010 0-6

≥0.010 0^*

[70]

Combination of transcription factors

MerR, CadR

Hg²⁺, Cd²⁺

egfp, mCherry

Escherichia coli TOP10

Detection of concurrent heavy metal contaminants in the environment

0-40 (Hg²⁺), 0-200 (Cd²⁺)

[71]

CadC, CadR

Cd²⁺

egfp and mCherry

Escherichia coli TOP10

Detection of high concentrations of biologically available Cd in water

≥0.050 0 (CadC-eGFP), ≥0.100 0 (CadR-mCherry)

[72]

MerR, PbrR

Hg²⁺, Pb²⁺

Indigoidine biosynthetic module

Escherichia coli TOP10

Detection of Hg²⁺ and Pb²⁺ in environmental samples

≥0.008 0

[73]

CadR, MerR

Cd²⁺, Pb²⁺, Hg²⁺

vioABE, vioC

Escherichia coli TOP10

Designed for detecting heavy metal contaminants in seawater

0.004 9-40, ≥0.004 9 (Cd²⁺), 0.024 4-200, ≥0.024 4 (Pb²⁺), 0.003 7-0.468 8, ≥0.000 5 (Hg²⁺)^*

[74]

UzcRS, UrpRS

UO₂²⁺

gfp

Caulobacter vibrioides

Detection of uranium (U) in groundwater samples

≥1

[75-76]

Sensing element	Detection object	Optimization strategy	Application	Performance enhancement (μmol/L)	References
ZntR	Cr²⁺ or Pb²⁺	Protein engineering	Cr and Pb-responsive WCMB were developed from the znt-manipulation subsystem for the determination of PB²⁺ or Cr²⁺ in environmental system	New selectivity for Cr²⁺ or Pb²⁺	[90]
	Pb²⁺	Protein engineering	The content of Pb²⁺ in environmental samples was monitored	0-0.010 0, <0.005 0^*	[58]
ArsR	Pb²⁺	Protein engineering	The content of Pb²⁺ in environmental samples was monitored	Detection accuracy>95%	[91]
MerR	Hg²⁺	Protein engineering	The content of Hg²⁺ in water samples was monitored	≥0.020 4	[92]
	Hg²⁺ Zn²⁺ Cu²⁺	Protein engineering	A new chimeric regulator WCMB was developed by replacing the metal-binding domain of MerR with ZntR or CueR	≥0.001 0 Hg²⁺ (MerR-Luc) ≥30 Zn²⁺ (MerRZntR-Luc) ≥10 Cu²⁺ (MerRCueR-Luc)	[93]
CadR	Cd²⁺	Protein engineering	The content of Cd²⁺ in water samples was monitored	≥0.450 0 μg/L	[94]
	Cd²⁺	Protein engineering	The interference of Hg²⁺ was reduced and the response to Cd²⁺ was enhanced	0.500 0-100， ≥0.079 0^*	[30]
MerR	Hg²⁺	Protein engineering	The “Parabola principle” is proposed and a visualized WCMB for Hg²⁺ detection in natural water is developed	0.200 0-0.250 0	[95]
	Hg²⁺	Protein engineering Promoter engineering	A super-sensitive visualized WCMB for the detection of ultra-trace Hg²⁺ was developed	≥0.313 0ng/L. When the fluorescence signal is more than 2.500 0 ng/l, it can be observed directly^*	[96]
ArsR	AsO₃^3-	Promoter engineering	A high sensitivity AsO₃^3- WCMB was developed, and the background reduction and signal output improvement were realized	0.100 0-4, ≥0.010 0	[97]
	AsO₃^3-	Promoter engineering	A highly sensitive and specific WCMB for AsO₃^3- detection in drinking water was developed	≥0.010 0^*	[98]
	AsO₃^3-	Promoter engineering	A functional promoter library screening method was developed, and different AsO₃^3- responsive elements arsR and OsmE1 were obtained, and a novel OsmE1 biosensor was constructed	≥0.040 0	[99]
	AsO₃^3-	Promoter engineering	A highly sensitive and specific WCMB for AsO₃^3- detection in drinking water was developed	≥0.100 0	[100]
ArsR MerR	AsO₃^3- Hg²⁺	Promoter engineering	A modular cascaded signal amplification method was developed, and a low-cost, portable and accurate AsO₃^3- and Hg²⁺ ultra-sensitive WCMB was constructed	≥0.100 0 ppb AsO₃^3- ≥0.010 0 ppb Hg²⁺	[101]
CadR	Cd²⁺	Promoter engineering	A highly sensitive and specific Cd²⁺ responsive WCMB was developed for the detection of Cd²⁺ in water	≥0.010 0	[102]
	Cd²⁺	Promoter engineering	A Cd²⁺ responsive WCMB with a negative feedback amplifier was developed to detect Cd²⁺ below the WHO and FAO standards with extremely high sensitivity and specificity	≥0.000 1^*	[103]

Sensing element

Detection object

Optimization strategy

Application

Performance enhancement

(μmol/L)

References

ZntR

Cr²⁺ or Pb²⁺

Protein engineering

Cr and Pb-responsive WCMB were developed from the znt-manipulation subsystem for the determination of PB²⁺ or Cr²⁺ in environmental system

New selectivity for Cr²⁺ or Pb²⁺

[90]

Pb²⁺

Protein engineering

The content of Pb²⁺ in environmental samples was monitored

0-0.010 0, <0.005 0^*

[58]

ArsR

Pb²⁺

Protein engineering

The content of Pb²⁺ in environmental samples was monitored

Detection accuracy>95%

[91]

MerR

Hg²⁺

Protein engineering

The content of Hg²⁺ in water samples was monitored

≥0.020 4

[92]

Hg²⁺

Zn²⁺

Cu²⁺

Protein engineering

A new chimeric regulator WCMB was developed by replacing the metal-binding domain of MerR with ZntR or CueR

≥0.001 0 Hg²⁺ (MerR-Luc)

≥30 Zn²⁺ (MerRZntR-Luc)

≥10 Cu²⁺ (MerRCueR-Luc)

[93]

CadR

Cd²⁺

Protein engineering

The content of Cd²⁺ in water samples was monitored

≥0.450 0 μg/L

[94]

Cd²⁺

Protein engineering

The interference of Hg²⁺ was reduced and the response to Cd²⁺ was enhanced

0.500 0-100， ≥0.079 0^*

[30]

MerR

Hg²⁺

Protein engineering

The “Parabola principle” is proposed and a visualized WCMB for Hg²⁺ detection in natural water is developed

0.200 0-0.250 0

[95]

Hg²⁺

Protein engineering

Promoter engineering

A super-sensitive visualized WCMB for the detection of ultra-trace Hg²⁺ was developed

≥0.313 0ng/L. When the fluorescence signal is more than 2.500 0 ng/l, it can be observed directly^*

[96]

ArsR

AsO₃^3-

Promoter engineering

A high sensitivity AsO₃^3- WCMB was developed, and the background reduction and signal output improvement were realized

0.100 0-4, ≥0.010 0

[97]

AsO₃^3-

Promoter engineering

A highly sensitive and specific WCMB for AsO₃^3- detection in drinking water was developed

≥0.010 0^*

[98]

AsO₃^3-

Promoter engineering

A functional promoter library screening method was developed, and different AsO₃^3- responsive elements arsR and OsmE1 were obtained, and a novel OsmE1 biosensor was constructed

≥0.040 0

[99]

AsO₃^3-

Promoter engineering

A highly sensitive and specific WCMB for AsO₃^3- detection in drinking water was developed

≥0.100 0

[100]

ArsR

MerR

AsO₃^3-

Hg²⁺

Promoter engineering

A modular cascaded signal amplification method was developed, and a low-cost, portable and accurate AsO₃^3- and Hg²⁺ ultra-sensitive WCMB was constructed

≥0.100 0 ppb AsO₃^3-

≥0.010 0 ppb Hg²⁺

[101]

CadR

Cd²⁺

Promoter engineering

A highly sensitive and specific Cd²⁺ responsive WCMB was developed for the detection of Cd²⁺ in water

≥0.010 0

[102]

Cd²⁺

Promoter engineering

A Cd²⁺ responsive WCMB with a negative feedback amplifier was developed to detect Cd²⁺ below the WHO and FAO standards with extremely high sensitivity and specificity

≥0.000 1^*

[103]