To fully understand enzymatic dynamics, it is essential to explore the complete conformational space of a biological catalyst. The catalytic mechanism of the nickel-dependent urease, the most efficient enzyme known, holds significant relevance for medical, pharmaceutical, and agro-environmental applications. A critical aspect of urease function is the conformational change of a helix-turn-helix motif that covers the active site cavity, known as the mobile flap. This motif has been observed in either an open or a closed conformation through X-ray crystallography studies and has been proposed to stabilize the coordination of a urea molecule to the essential dinuclear Ni(II) cluster in the active site, a requisite for subsequent substrate hydrolysis. This study employs cryo-electron microscopy (cryo-EM) to investigate the transient states within the conformational space of the mobile flap, devoid of the possible constraints of crystallization conditions and solid-state effects. By comparing two cryo-EM structures of Sporosarcina pasteurii urease, one in its native form and the other inhibited by N-(n-butyl) phosphoric triamide (NBPTO), we have identified an intermediate state between the open and the catalytically efficient closed conformation of the helix-turn-helix motif, suggesting a role of its tip region in this transition between the two states.1
[1] L. Mazzei; G. Tria; S. Ciurli; M. Cianci Int. J. Biol. Macromol. 2024, 283, 137904