Genetic circuit-enabled synthetic biosensors： design and applications Synthetic Biology Journal    DOI: 10.12211/2096-8280.2025-031

Fig. 10 Examples of improving the response speed of genetic circuit-enabled synthetic biosensors
(a) An E. coli-based sensor with a synthetic electron transfer chain that express an electrical protein switch enable rapid detection of an endocrine disruptor 4-hydroxytamoxifen (4-HT)^[6]; (b) Reversible response kinetics curve and sensing mechanism of a protein phosphorylation signaling network with rapid activation and deactivation^[197].

Extracts from the Article

当前，基于基因线路的合成生物传感器在响应速度方面仍面临巨大挑战。相较于已实现实时检测的电子或化学传感技术，依赖基因转录-翻译级联反应的传感器通常需要30分钟至数小时才能输出信号，这一滞后性严重制约其产业化应用进程［167］。近年来，研究者通过创新传感机制已取得突破性进展。Atkinson等［6］构建了基于合成电子传递链的电流输出型生物传感器，可于2分钟内感知硫代硫酸盐，暴露于内分泌干扰物（4-HT）环境中3分钟内产生可检测的电流输出（图10a）；Yang等［197］在哺乳动物细胞中设计了快速激活/失活的磷酸化信号网络，该体系感知肿瘤坏死因子TNF-α并动态调控IL-10分泌过程，实现了相较于传统哺乳细胞传感方式更加快速的可逆性响应（图10b）。在蛋白质工程领域，翻译后修饰、构象转换及FRET（F?rster Resonance Energy Transfer）技术为快速传感提供了核心策略。Wang等［198］开发的蛋白酶修饰快速分泌系统（PASS）创新性地利用内质网驻留信号预存目标蛋白，在检测到小分子、光或抗原时将分裂的切割酶二聚化以激活其切割功能，实现了治疗性蛋白的快速可控释放；Torres等［199］借助深度学习设计出可特异性结合螺旋肽的人工变构蛋白，据此构建了变构发光型的快速生物传感器；在FRET技术领域，Hellweg等［200］构建了模块化FRET传感器构建平台ChemoX，通过替换荧光蛋白或合成荧光团可实现多色信号输出，已成功开发出钙离子、ATP等靶标的分钟级响应传感器。此外，本文2.3部分所述传感器多模态输出模块中的光-电信号转换［48，166］、比色传感［42，160-161］和空间图案传感［32，36］等系统可使传感输出信号以方便识读的形式快速获取，加速传感器信号报告。这些创新机制（包括高效信号级联、预存元件激活、变构效应利用等）和创新报告形式为突破合成生物传感器响应速度瓶颈提供了新思路。未来研究需通过开发新型信号识别机制与信号报告策略，构建兼具快速响应与精准识别的新一代生物传感器。

Other Images/Table from this Article

• Table 1  Typical genetic circuit-enabled synthetic biosensors and applications
• Fig. 1  Mechanisms and applications of transcriptional regulatory protein-enabled synthetic biosensors(a-c) Mechanisms of allosteric transcription factors, transmembrane transcription factors, and two-component systems-enabled biosensors; (d) Response curves of biosensors based on activation-type and repression-type transcription factor; (e) Gene circuit of a thiosulfate-sensing sensor based on the two-component system ThsRS^[54]; (f) Gene circuit of a transcription factor-enabled biosensor for monitoring glucose uptake^[14]; (g) Gene circuit of a target-dependent RNA polymerase-enabled biosensor^[95].
• Fig. 2  Mechanisms of riboswitches-enabled synthetic biosensors(a) Riboswitch-enabled sensors based on transcriptional activation and transcriptional repression.(b) Riboswitch-enabled sensors based on translational activation and translational repression..
• Fig. 3  Mechanisms and applications of riboregulator-enabled synthetic biosensors(a) Mechanism of the toehold switch^[124]; (b) A ribozyme-based sensing platform RENDR^[125]; (c) Gene circuit of a sensor for reporting bacterial DNA transfer in soil using split ribozymes^[133]; (d) Mechanism of the group I intron-enabled RNA trans-splicing system SENTR and its application for intracellular mRNA detection ^[122].
• Fig. 4  Mechanisms of CRISPR-enabled synthetic biosensors(a) RNA sensing enabled by reprogrammed tracrRNA in type II CRISPR system^[17]; (b) RNA sensing enabled by RNA-triggered RNA processing^[142]; (c) RNA sensing enabled by strand displacement reactions^[143].
• Table 2  Advantages and limitations of genetic circuit-enabled synthetic biosensors based on different signal recognition mechanisms
• Fig. 5  Typical biomemory modules for recording biosensing signals(a) Gene circuit for a biomemory module based on toggle switch; (b) Gene circuit of a zinc ion memory module based on the recombinase flip mechanism^[153]; (c) Principle of the CAMERA recording system based on CRISPR/Cas^[158]; (d) Gene circuit of the DNA sensing and recording module (CATCH) based on CRISPR/Cas^[159].
• Fig. 6  Circuit designs and applications of multilayered biocomputing modules(a) Gene circuit of a dual-parameter biosensor based on an AND logic gate and its response heatmap^[104]; (b) Gene circuit of a metabolic control sensor integrating a multi-input genetic logic circuit ^[163];
• Fig. 7  Synthetic biosensing systems with multimodal outputs for integration with various detection platforms(a) A agarose hydrogel-encapsulated arsenic ion sensor for visual array display on a smartphone platform^[32]; (b) A sensor that visualizes biosensing signals through the formation of bacterial spatial motility patterns^[36]; (c) A sensing system coupling a fluorescent sensor with a photoelectric chip to convert optical signals into wireless signals^[48].
• Fig. 8  Strategies for increasing the sensitivity of genetic circuit-enabled synthetic biosensors(a) Typical response curves and characteristics of genetic circuit-enabled synthetic biosensors.; (b) Dynamic range optimization based on promoter engineering; (c) Limit of detection optimization by gradient regulation of biorecognition components concentration and target enrichment technology; (d) Dynamic range optimization using sensing signal amplifiers based on activator transcription factors, positive feedback loops and cascade repressors ^[7]; (e) Leakage optimization utilizing antisense RNA-mediated translation blocking, decoy DNA binding site competitive inhibition, and output protein degradation tags^[1].
• Fig. 9  Strategies for increasing the specificity of genetic circuit-enabled synthetic biosensors(a) Semi-rational design and saturation mutagenesis of the transcription factor LysG to prevent its binding to L-lysine (L-Lys) while retaining its ability to bind L-arginine (L-Arg) and L-histidine (L-His)^[191]; (b) Semi-rational design and saturation mutagenesis of the TynA-FeaR sensing system for specific detection of phenylethylamine. (DA: dopamine; PEA: phenylethylamine; Tyra: tyramine; Trypta: tryptamine)^[192]; (c) Coupling transcription factors ZraR and ZntR through an AND gate for specific respond to Zn²⁺^[7,21].
• Fig. 11  Examples of stability and biosafety optimization for genetic circuit-enabled synthetic biosensors(a) A plasmid copy number control platform in Escherichia coli based on dynamic regulation of translation initiation factors^[204]; (b) A biocontaining CRISPR-Cas9 dual-modal kill switch (CRISPRks) for E. coli under the dual control of chemical inducer anhydrotetracycline (aTc) and temperature changes^[212].