title: A Fluorescent, Reagentless Biosensor for ATP, Based on Malonyl-Coenzyme A Synthetase.
abstract: 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.
authors.affiliation: The Francis Crick Institute , Mill Hill Laboratory, The Ridgeway, Mill Hill, London NW7 1AA, United Kingdom.
authors.forename: Renée
authors.initials: R
authors.lastname: Vancraenenbroeck
authors.affiliation: The Francis Crick Institute , Mill Hill Laboratory, The Ridgeway, Mill Hill, London NW7 1AA, United Kingdom.
authors.forename: Martin R
authors.initials: MR
authors.lastname: Webb
mesh_terms: D000255:Adenosine Triphosphate / Q000032:analysis* / Q000737:chemistry; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism; D015374:Biosensing Techniques; D003066:Coenzyme A Ligases / Q000737:chemistry* / Q000378:metabolism; D005456:Fluorescent Dyes / Q000737:chemistry*; D008954:Models, Biological; D011770:Pyruvate Kinase / Q000737:chemistry / Q000494:pharmacology
chemical_list: D001426:Bacterial Proteins; D005456:Fluorescent Dyes; D000255:Adenosine Triphosphate; D011770:Pyruvate Kinase; D003066:Coenzyme A Ligases; C021533:malonyl-CoA synthetase