A Selective Fluorescent l-Lactate Biosensor Based on an l-Lactate-Specific Transcription Regulator and Förster Resonance Energy Transfer.
A Selective Fluorescent l-Lactate Biosensor Based on an l-Lactate-Specific Transcription Regulator and Förster Resonance Energy Transfer.
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DOI：
10.3390/bios12121111
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发表时间：
2022-12-01
期刊：
影响因子：
5.4
通讯作者：
Ma, Cuiqing
中科院分区：
文献类型：
--
作者：
Xu, Xianzhi;Xu, Rong;Hou, Shuang;Kang, Zhaoqi;Lue, Chuanjuan;Wang, Qian;Zhang, Wen;Wang, Xia;Xu, Ping;Gao, Chao;Ma, Cuiqing
关键词：
食品、临床和细菌发酵样品中l-乳酸水平的选择性检测引起了广泛关注。许多基于非立体选择性识别元件的荧光生物传感器已被开发用于乳酸检测。本文中，来自沙门氏菌鼠伤寒血清型LT 2的变构转录因子STLldR被鉴定为对l-乳酸立体选择性应答。然后，STLldR与Förster共振能量转移（FRET）相结合，构建了荧光l-乳酸生物传感器FILLac。FILac通过在青色和黄色荧光蛋白之间截短STLldR的N-和C-末端氨基酸来进一步优化。优化后的生物传感器FILLac 10 N 0 C的最大发射比变化（ΔRmax）为33.47 ± 1.91%，表观解离常数（Kd）为6.33 ± 0.79 μM，检测限为0.68 μM。将FILLac 10 N 0 C应用于96孔微孔板中，检测细菌发酵样品和商业食品（如胶苏和酸奶）中的l-乳酸。FILLac 10 N 0 C的定量结果与商业L-乳酸生物传感器SBA-40 D生物分析仪的定量结果表现出良好的一致性。因此，与高通量检测兼容的生物传感器FILLac 10 N 0 C可能是用于定量不同生物样品中l-乳酸的潜在选择。
Selective detection of l-lactate levels in foods, clinical, and bacterial fermentation samples has drawn intensive attention. Many fluorescent biosensors based on non-stereoselective recognition elements have been developed for lactate detection. Herein, the allosteric transcription factor STLldR from Salmonella enterica serovar Typhimurium LT2 was identified to be stereo-selectively respond to l-lactate. Then, STLldR was combined with Förster resonance energy transfer (FRET) to construct a fluorescent l-lactate biosensor FILLac. FILLac was further optimized by truncating the N- and C-terminal amino acids of STLldR between cyan and yellow fluorescent proteins. The optimized biosensor FILLac10N0C exhibited a maximum emission ratio change (ΔRmax) of 33.47 ± 1.91%, an apparent dissociation constant (Kd) of 6.33 ± 0.79 μM, and a limit of detection of 0.68 μM. FILLac10N0C was applied in 96-well microplates to detect l-lactate in bacterial fermentation samples and commercial foods such as Jiaosu and yogurt. The quantitation results of FILLac10N0C exhibited good agreement with that of a commercial l-lactate biosensor SBA-40D bioanalyzer. Thus, the biosensor FILLac10N0C compatible with high-throughput detection may be a potential choice for quantitation of l-lactate in different biological samples.
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影响因子：
4.6
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