Dynamics in Thermotoga neapolitana Adenylate Kinase: 15N Relaxation and Hydrogen−Deuterium Exchange Studies of a Hyperthermophilic Enzyme Highly Active at 30 °C†

Backbone conformational dynamics of Thermotoga neapolitana adenylate kinase in the free form (TNAK) and inhibitor-bound form (TNAK*Ap5A) were investigated at 30 °C using 15N NMR relaxation measurements and NMR monitored hydrogen−deuterium exchange. With kinetic parameters identical to those of Escherichia coli AK (ECAK) at 30 °C, TNAK is a unique hyperthermophilic enzyme. These catalytic properties make TNAK an interesting and novel model to study the interplay between protein rigidity, stability, and activity. Comparison of fast time scale dynamics (picosecond to nanosecond) in the open and closed states of TNAK and ECAK at 30 °C reveals a uniformly higher rigidity across all domains of TNAK. Within this framework of a rigid TNAK structure, several residues located in the AMP-binding domain and in the core−lid hinge regions display high picosecond to nanosecond time scale flexibility. Together with the recent comparison of ECAK dynamics with those of hyperthermophilic Aquifex aeolicus AK (AAAK), our resu...