"Thermodynamic Analysis Of Atp Dissociation In A Dead-Box Protein"

RNA plays an essential role in many cellular functions. Similar to proteins, RNA function is strictly determined by the overall structure and dynamics of the RNA macromolecule. Thus, many cellular proteins or accessory factors are associated with RNA folding. One example of these RNA binding proteins are DEAD‐Box proteins. DEAD‐Box proteins utilize ATP hydrolysis to unwind and reanneal RNA duplexes to promote correct folding during RNA processing. DEAD‐Box proteins possess two conserved RecA‐like domains that bind ATP and RNA. In addition, these proteins also contain N‐ and/or C‐ terminal domains (NTD and CTD, respectively). Peripheral domains in DEAD‐Box proteins are proposed to regulate ATP hydrolysis as well as confer RNA specificity. Thus, the NTD and/or CTD can alter the ATP binding pocket and subsequently alter ATP association and/or dissociation. For example, Rok1p, a yeast DEAD‐Box protein, contains both a NTD and a CTD. The NTD in Rok1p is suggested to affect the structure and stability of the protein. Therefore, the dissociation rate of a fluorescence ATP analog was determined through transient kinetics and stopped‐flow spectroscopy under various thermal conditions. Resulting trends conform to double exponential fits suggesting a two‐step mechanism. The observed slow rate was consistent with a conformation change in the protein that allows rapid dissociation. The rate in which the conformation change occurs was observed to be temperature dependent. The current hypothesis centers on the ability, of the Rok1p peripheral domains, to affect the slow structural change needed for ATP dissociation.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.