Quantification of riboflavin, flavin mononucleotide, and flavin adenine dinucleotide in mammalian model cells by CE with LED‐induced fluorescence detection

黄素组 黄素单核苷酸 黄素腺嘌呤二核苷酸 核黄素 辅因子 黄蛋白 荧光 生物化学 化学 氧化还原 生物物理学 生物 量子力学 物理 有机化学
作者
Jens Hühner,Álvaro Inglés‐Prieto,Christian Neusüß,Michael Lämmerhofer,Harald Janovjak
出处
期刊:Electrophoresis [Wiley]
卷期号:36 (4): 518-525 被引量:50
标识
DOI:10.1002/elps.201400451
摘要

Cultured mammalian cells essential are model systems in basic biology research, production platforms of proteins for medical use, and testbeds in synthetic biology. Flavin cofactors, in particular flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD), are critical for cellular redox reactions and sense light in naturally occurring photoreceptors and optogenetic tools. Here, we quantified flavin contents of commonly used mammalian cell lines. We first compared three procedures for extraction of free and noncovalently protein‐bound flavins and verified extraction using fluorescence spectroscopy. For separation, two CE methods with different BGEs were established, and detection was performed by LED‐induced fluorescence with limit of detections (LODs 0.5–3.8 nM). We found that riboflavin (RF), FMN, and FAD contents varied significantly between cell lines. RF (3.1–14 amol/cell) and FAD (2.2–17.0 amol/cell) were the predominant flavins, while FMN (0.46–3.4 amol/cell) was found at markedly lower levels. Observed flavin contents agree with those previously extracted from mammalian tissues, yet reduced forms of RF were detected that were not described previously. Quantification of flavins in mammalian cell lines will allow a better understanding of cellular redox reactions and optogenetic tools.
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