YnLac(炔基乳酸钠)|安捷凯

YnLac(炔基乳酸钠)

YnLac是一种生物正交化学探针，是L型乳酸的炔基衍生物，主要用于乳酸代谢研究，可以与叠氮荧光试剂发生点击反应click），作为哺乳动物细胞中蛋白乳酸化修饰的蛋白质组学层面的检测和鉴定，也可与叠氮生物素（Biotin）点击用于蛋白质组学中的富集与鉴定等。

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Background

L-Lactate, the end product of glycolysis generated from the reduction of pyruvate by lactate dehydrogenase, is an essential metabolite in mammals. Although historically considered to be a metabolic waste product, L-lactate has since been recognized to serve as a major circulating energy source and, additionally, act as a multifunctional signaling molecule involved in diverse physiological and pathological processes, including angiogenesis,inflammation, neural activity, neoplasia, and autoimmune diseases.Recently, a seminal study by Zhao laboratory reported that L-lactate is a precursor to drive direct modifications of histones.¹

sodium (S)-2-hydroxypent-4-ynoate (YnLac), that contains a terminal alkynyl on the original methyl group , as a potential chemical reporter for the detection and profiling of protein lactylation.¹

Protocol

HeLa cells were cultured on sterilized coverslips and incubated with YnLac (20 mM, 2 M stock solution in PBS) in culture medium. The same volume of PBS was used as the vehicle control. After 12 h of labeling at 37°C, the cells were washed once with warmed PBS, fixed with 4% formaldehyde in PBS for 10 min at room temperature, and then washed twice with ice-cold PBS. Cells were permeabilized with 0.3% Triton X-100 in PBS for 10 min at room temperature, blocked with 3% BSA in PBS for 30min at room temperature, and washed with PBS (3 x 5 min with gentle agitation). The cells were then treated with a freshly premixed click reaction cocktail (50 μM azido-rhodamine, 1 mM TCEP, 100 μM TBTA, and 1 mM CuSO4×5H2O) in PBST (0.1% Tween-20 in PBS) for 1 h at room temperature. After gentle washes three times with 1% Tween-20 in PBS, cells were then stained with Hoechst 33342 and imaged on a Nikon A1R confocal fluorescence microscope. For Hoechst channel, the 405 nm laser was used as the excitation, and emission was collected between 425 nm to 475 nm. For rhodamine channel, the 561 nm laser was used as the excitation, and emission was collected between 570 nm to 620 nm.¹

参考文献：

1.Yanan Sun, Yanchi Chen, and Tao Peng.A Bioorthogonal Chemical Reporter for the Detection and Identification of Protein Lactylation.Electronic Supplementary Material (ESI) for Chemical Science. This journal is © The Royal Society of Chemistry 2022

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生命科学

有机化学

无机化学

材料科学

YnLac(炔基乳酸钠)

YnLac(炔基乳酸钠)详情

Background

Protocol

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