Vancraenenbroeck Renée, Webb Martin R
The Francis Crick Institute , Mill Hill Laboratory, The Ridgeway, Mill Hill, London NW7 1AA, United Kingdom.
ACS Chem Biol. 2015 Nov 20;10(11):2650-7. doi: 10.1021/acschembio.5b00346. Epub 2015 Sep 21.
A fluorescent reagentless biosensor for ATP has been developed, based on malonyl-coenzyme A synthetase from Rhodopseudomonas palustris as the protein scaffold and recognition element. Two 5-iodoacetamidotetramethylrhodamines were covalently bound to this protein to provide the readout. This adduct couples ATP binding to a 3.7-fold increase in fluorescence intensity with excitation at 553 nm and emission at 575 nm. It measures ATP concentrations with micromolar sensitivity and is highly selective for ATP relative to ADP. Its ability to monitor enzymatic ATP production or depletion was demonstrated in steady-state kinetic assays in which ATP is a product or substrate, respectively.
一种用于ATP的无荧光试剂生物传感器已被开发出来，它以沼泽红假单胞菌的丙二酰辅酶A合成酶作为蛋白质支架和识别元件。两个5-碘乙酰胺基四甲基罗丹明共价结合到该蛋白质上以提供读数。这种加合物将ATP结合与在553nm激发和575nm发射时荧光强度增加3.7倍联系起来。它以微摩尔灵敏度测量ATP浓度，并且相对于ADP对ATP具有高度选择性。在稳态动力学测定中分别证明了其监测酶促ATP产生或消耗的能力，其中ATP分别是产物或底物。
