Unexpected Sophistication and Complexity of Chemomechanical Control Linkages in Nucleic Acid Motor

Abstract

Ring-shaped, oligomeric ATPases are essential for a variety of cellular processes ranging from protein and nucleic acid metabolism to organelle transport. A subset of these motor proteins, the hexameric helicases, couple the binding and hydrolysis of ATP to the physical manipulation of nucleic acids to support essential cellular processes such as gene regulation, DNA replication, and DNA repair. How nucleotide turnover is coordinated between six independent motor subunits to generate helicase movement has been a long-standing question in the field. The topological problem of how extended nucleic substrates are loaded into the central pore of a closed hexameric toroid is similarly not well-understood.Using the 300 kDa E. coli Rho protein as a model hexameric RNA/DNA helicase, we have examined these issues through a combined structural and biochemical approach. Models derived from high-resolution crystallographic and solution X-ray scattering data explain in molecular detail how the Rho ring opens and closes in response to substrate binding, how the ATP cycle is coordinated with RNA movement through the motor interior, and why certain helicase families translocate with a preferred directional polarity. We find an unexpectedly rich array of physical connectors that sense the RNA binding status of individual subunits in the motor, and that appear to couple these states to the timing of ATP hydrolysis in a manner that is responsive to the base sequence of the substrate. These studies highlight the physical complexity of hexameric helicases, and suggest that related motor proteins utilize similarly intricate chemomechanical linkage mechanisms.