![]() Similarly, in lower resolution X-ray structures (>2.8 Å) AlF 3 cannot be distinguished unambiguously from AlF 4 − 9, 10. distinction between AlF 3 and MgF 3 −, has been raised and it could be shown by 19F NMR spectroscopy that some complexes, which were believed to contain AlF 3, contain instead MgF 3 − 9. However, some concern regarding their discrimination, e.g. The formation of AlF 4 − or AlF 3 is controlled by pH, the latter being favored at lower pH values 9. AlF 4 − forms together with the phosphate oxygen atom of ADP as well as an apical water molecule an octahedral complex mimicking the “in-line” anionic transition state of phosphoryl transfer, whereas AlF 3 and MgF 3 − form trigonal-bipyramidal complexes 7. The number of deposited protein structures containing analogues such as AlF 4 −, AlF 3 and MgF 3 − has strongly increased in the past years 7. Metal fluorides have been found to mimic such states for structural studies, mostly using X-ray crystallography 7, 8. 1 for a sketch of the limiting case of an associative ATP hydrolysis mechanism 6). Of particular interest in unravelling the ATP hydrolysis reaction mechanism is the transition state of the phosphoryl (PO 3 −) transfer reaction (see Fig. ![]() Diverse ATP analogues can be employed to mimic different stages of ATP hydrolysis as closely as possible 3, 4 which, in combination with molecular dynamics simulations 5, can give mechanistic insights into complex biomolecular reaction coordinates. Structural insights into the functioning of these molecular machines are not straightforward to obtain, neither by X-ray crystallography, nor by cryo-electron microscopy or NMR spectroscopy due to the difficulty in trapping the intermediate catalytic states occurring during ATP hydrolysis. For example, motor proteins belong to the class of ATPases, which hydrolyze ATP into ADP (adenosine diphosphate) and inorganic phosphate to gain chemical energy allowing such enzymes to drive further chemical or mechanical events 2. 19F and 27Al MAS NMR spectra reveal a highly mobile, fast-rotating aluminum fluoride unit pointing to the capture of a late ATP hydrolysis transition state in which the phosphoryl unit is already detached from the arginine and lysine fingers.Īdenosine triphosphate (ATP)-driven motor proteins play a key role in various cellular processes 1. We determined the positioning of the metal ion cofactor within the active site using electron paramagnetic resonance, and identified the protein protons coordinating to the phosphate groups of ADP and DNA using proton-detected 31P, 1H solid-state nuclear magnetic resonance spectroscopy at fast magic-angle spinning > 100 kHz, as well as temperature-dependent proton chemical-shift values to prove their engagements in hydrogen bonds. ![]() We present atomic-level structural and dynamic insights on a state created by ADP aluminum fluoride binding to the bacterial DnaB helicase from Helicobacter pylori. The ATP hydrolysis transition state of motor proteins is a weakly populated protein state that can be stabilized and investigated by replacing ATP with chemical mimics.
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