Magnesium induced structural reorganization in the active site of adenylate kinase.

Phosphoryl transfer is a fundamental reaction in cellular signaling and metabolism that requires Mg as an essential cofactor. While the primary function of Mg is electrostatic activation of substrates, such as ATP, the full spectrum of catalytic mechanisms exerted by Mg is not known. In this study, we integrate structural biology methods, molecular dynamic (MD) simulations, phylogeny, and enzymology assays to provide molecular insights into Mg-dependent structural reorganization in the active site of the metabolic enzyme adenylate kinase.

Insights into Enzymatic Catalysis from Binding and Hydrolysis of Diadenosine Tetraphosphate by . Adenylate Kinase.

Adenylate kinases play a crucial role in cellular energy homeostasis through the interconversion of ATP, AMP, and ADP in all living organisms. Here, we explore how adenylate kinase (AdK) from interacts with diadenosine tetraphosphate (AP4A), a putative alarmone associated with transcriptional regulation, stress, and DNA damage response. From a combination of EPR and NMR spectroscopy together with X-ray crystallography, we found that AdK interacts with AP4A with two distinct modes that occur on disparate time scales.

Site-directed attachment of photoexcitable spin labels for light-induced pulsed dipolar spectroscopy.

Photoexcited triplet states are gaining popularity as spin labels in pulsed electron paramagnetic resonance (EPR) spectroscopy. Here, we demonstrate that the fluorophores Eosin Y, Rose Bengal and Atto Thio12 are suitable markers for distance determination by laser-induced magnetic dipole (LaserIMD) spectroscopy in proteins that lack an intrinsic photoexcitable center.

Optimising broadband pulses for DEER depends on concentration and distance range of interest.

EPR distance determination in the nanometre region has become an important tool for studying the structure and interaction of macromolecules. Arbitrary waveform generators (AWGs), which have recently become commercially available for EPR spectrometers, have the potential to increase the sensitivity of the most common technique, double electron-electron resonance (DEER, also called PELDOR), as they allow the generation of broadband pulses. There are several families of broadband pulses, which are different in general pulse shape and the parameters that define them.