Coupling of ATPase activity, microtubule binding, and mechanics in the dynein motor domain. (31st May 2019)

• Record Type: Journal Article
• Authors: Niekamp, Stefan; Coudray, Nicolas; Zhang, Nan; Vale, Ronald D; Bhabha, Gira
• Is Part Of: EMBO journal. Volume 38:Number 13(2019)
• Page Start: n/a
• Page End: n/a
• Publication Date: 2019-05-31
• Subjects: cryo‐electron microscopy -- dynein -- microtubule -- motility -- motor proteins; Molecular biology -- Periodicals; 572.805
• DOI: 10.15252/embj.2018101414
• Languages: English
• ISSNs: 0261-4189

Abstract

Abstract: The movement of a molecular motor protein along a cytoskeletal track requires communication between enzymatic, polymer‐binding, and mechanical elements. Such communication is particularly complex and not well understood in the dynein motor, an ATPase that is comprised of a ring of six AAA domains, a large mechanical element (linker) spanning over the ring, and a microtubule‐binding domain (MTBD) that is separated from the AAA ring by a ~ 135 Å coiled‐coil stalk. We identified mutations in the stalk that disrupt directional motion, have microtubule‐independent hyperactive ATPase activity, and nucleotide‐independent low affinity for microtubules. Cryo‐electron microscopy structures of a mutant that uncouples ATPase activity from directional movement reveal that nucleotide‐dependent conformational changes occur normally in one‐half of the AAA ring, but are disrupted in the other half. The large‐scale linker conformational change observed in the wild‐type protein is also inhibited, revealing that this conformational change is not required for ATP hydrolysis. These results demonstrate an essential role of the stalk in regulating motor activity and coupling conformational changes across the two halves of the AAA ring. Synopsis: The movement of dynein on microtubules requires communication between enzymatic, polymer‐binding, and mechanical elements. Our results reveal how dynein's coiled‐coil stalk plays a critical role in coordinating such domain movements. Stalk mutants disrupt unidirectional motion along microtubules, show nucleotide‐independent low affinity for microtubules, and lack microtubule‐regulation of ATPase activity. Cryo‐electron microscopy structures of one mutant show that nucleotide‐dependent conformational changes are disrupted in one half of the AAA ring while the other half is minimally affected; the partial ring conformational change results in unregulated ATP hydrolysis and blocks the linker conformational change that drives motility. Our structural and functional results suggest a model for how the stalk domain modulates conformational changes around the AAA ring, which are initiated by nucleotide binding at dynein's main ATP site (AAA1). Abstract : Mutations in the dynein stalk demonstrate its role in coordinating conformational changes across the AAA ring of the motor domain, and in regulating microtubule‐dependent movement and ATPase activity.