Substrate Binding Specifically Modulates Domain Arrangements in Adenylate Kinase

Abstract

The enzyme adenylate kinase (ADK) features two substrate binding domains that undergo large-scale motions during catalysis. In the apo state, the enzyme preferentially adopts a globally open state with accessible binding sites. Binding of two substrate molecules (AMP + ATP or ADP + ADP) results in a closed domain conformation, allowing efficient phosphoryl-transfer catalysis. We employed molecular dynamics simulations to systematically investigate how the individual domain motions are modulated by the binding of substrates. Two-dimensional free-energy landscapes were calculated along the opening of the two flexible lid domains for apo and holo ADK as well as for all single natural substrates bound to one of the two binding sites of ADK. The simulations reveal a strong dependence of the conformational ensembles on type and binding position of the bound substrates and a nonsymmetric behavior of the lid domains. Altogether, the ensembles suggest that, upon initial substrate binding to the corresponding lid site, the opposing lid is maintained open and accessible for subsequent substrate binding. In contrast, ATP binding to the AMP-lid induces global domain closing, preventing further substrate binding to the ATP-lid site. This might constitute a mechanism by which the enzyme avoids the formation of a stable but enzymatically unproductive state.

Assessing the Interactions of Statins with Human Adenylate Kinase Isoenzyme 1: Fluorescence and Enzyme Kinetic Studies

“…Noncompetitive inhibitors are known to bind to the enzyme at allosteric sites (i.e., locations other than its active site). Adenylate kinase is a small monomeric enzyme consisting of CORE and LID domains and two substrate binding sites (NMPbd and NTPbd) that are specifically occupied by AMP and ATP, or alternatively by one ADP molecule each [ 70 ]. The LID domain and CORE domain with NTPbd form the ATP binding pocket, while the NMPbd and CORE domain form the AMP binding pocket [ 71 ].…”

Hypoxanthine-Guanine Phosphoribosyltransferase/adenylate Kinase From Zobellia galactanivorans: A Bifunctional Catalyst for the Synthesis of Nucleoside-5′-Mono-, Di- and Triphosphates

“…Regarding to the ADP synthesis, K M for AMP is 10-fold higher than K M for ATP. This significant difference in the affinity between AMP and ATP is probably due to a more unstable binding of AMP onto AMP binding site, whereas the ATP binding onto LID domain seems to be favored ( Zeller and Zacharias, 2015 ). Moreover, the turnover numbers are in compliance with the release form of products, where ADP liberation from AMP occurs through AMP binding site, and ADP liberation from ATP occurs through CORE domain ( Zeller and Zacharias, 2015 ).…”

“…This significant difference in the affinity between AMP and ATP is probably due to a more unstable binding of AMP onto AMP binding site, whereas the ATP binding onto LID domain seems to be favored ( Zeller and Zacharias, 2015 ). Moreover, the turnover numbers are in compliance with the release form of products, where ADP liberation from AMP occurs through AMP binding site, and ADP liberation from ATP occurs through CORE domain ( Zeller and Zacharias, 2015 ). Finally, the difference in the catalytic efficiency between AMP and ATP fits the model described by Ådén et al (2013) where AMP can bind to LID with a very low affinity generating an unproductive complex ( Whitford et al, 2007 ; Ådén et al, 2013 ; Zeller and Zacharias, 2015 ).…”

“…To explain these results, a homology model of ZgHGPRT/AMPK was built (Figure 3), and then it was superposed with HGXPRT from Thermus thermophilus HB8 complexed with IMP (TtHGXPRT, PDB id 3ACD) (Kanagawa et al, 2010;. As shown in Figure 11, hypoxanthine binding is stabilized by a network of hydrogen bonds between Asp 104, Lys 132 and Ile 154, and N1, exocyclic ATP binding onto LID domain seems to be favored (Zeller and Zacharias, 2015). Moreover, the turnover numbers are in compliance with the release form of products, where ADP liberation from AMP occurs through AMP binding site, and ADP liberation from ATP occurs through CORE domain (Zeller and Zacharias, 2015).…”

Use of Integrated Computational Approaches in the Search for New Therapeutic Agents

“…Various computational approaches have been developed to discover the collective motions associated with the dynamics of proteins, most of them applied to protein folding simulations and ligand‐protein docking simulations ,. The time scales of the conformational transitions of proteins are frequently of order of microseconds to seconds while MD is restricted to integration timesteps of a few femtoseconds.…”