A Genetically Encoded Fluorescent Biosensor for Intracellular Measurement of Malonyl-CoA

被引：0
作者：
Ranzau, Brodie L. [1 ]
Robinson, Tiffany D. [1 ]
Scully, Jack M. [1 ]
Kapelczak, Edmund D. [2 ]
Dean, Teagan S. [1 ]
Teslaa, Tara [2 ,3 ]
Schmitt, Danielle L. [1 ,3 ,4 ]
机构：
[1] Univ Calif Los Angeles, Dept Chem & Biochem, Los Angeles, CA 90095 USA
[2] Univ Calif Los Angeles, David Geffen Sch Med, Dept Mol & Med Pharmacol, Los Angeles, CA 90095 USA
[3] Univ Calif Los Angeles, Mol Biol Inst, Los Angeles, CA 90095 USA
[4] Univ Calif Los Angeles, Inst Quantitat & Computat Biosci, Los Angeles, CA 90095 USA
来源：
ACS BIO & MED CHEM AU | 2024年 / 5卷 / 01期
基金：
美国国家卫生研究院;
关键词：
malonyl-CoA; biosensor; fluorescence; metabolism; fatty acid biosynthesis; LYSINE MALONYLATION; BIOSYNTHESIS; METABOLISM;
D O I：
10.1021/acsbiomedchemau.4c00103
中图分类号：
Q5 [生物化学]; Q7 [分子生物学];
学科分类号：
071010 ; 081704 ;
摘要：
Malonyl-CoA is the essential building block of fatty acids and regulates cell function through protein malonylation and allosteric regulation of signaling networks. Accordingly, the production and use of malonyl-CoA is finely tuned by the cellular energy status. Most studies of malonyl-CoA dynamics rely on bulk approaches that take only a snapshot of the average metabolic state of a population of cells, missing out on heterogeneous differences in malonyl-CoA and fatty acid biosynthesis that could be occurring among a cell population. To overcome this limitation, we have developed a genetically encoded fluorescent protein-based biosensor for malonyl-CoA that can be used to capture malonyl-CoA dynamics in single cells. This biosensor, termed Malibu ( mal onyl-CoA i ntracellular b iosensor to u nderstand dynamics), exhibits an excitation-ratiometric change in response to malonyl-CoA binding. We first used Malibu to monitor malonyl-CoA dynamics during inhibition of fatty acid biosynthesis using cerulenin in Escherichia coli, observing an increase in Malibu response in a time- and dose-dependent manner. In HeLa cells, we used Malibu to monitor the impact of fatty acid biosynthesis inhibition on malonyl-CoA dynamics in single cells, finding that two inhibitors of fatty acid biosynthesis, cerulenin and orlistat, which inhibit different steps of fatty acid biosynthesis, increase malonyl-CoA levels. Altogether, we have developed a new genetically encoded biosensor for malonyl-CoA, which can be used to study malonyl-CoA dynamics in single cells, providing an unparalleled view into fatty acid biosynthesis.
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收藏
页码：184 / 193
页数：10
相关论文
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