Allosteric Communication in the Dynein Motor Domain

Abstract

SUMMARY Dyneins power microtubule motility using ring-shaped, AAA-containing motor domains. Here, we report X-ray and electron microscopy (EM) structures of yeast dynein bound to different ATP analogs, which collectively provide insight into the roles of dynein’s two major ATPase sites, AAA1 and AAA3, in the conformational change mechanism. ATP binding to AAA1 triggers a cascade of conformational changes that propagate to all six AAA domains and cause a large movement of the “linker,” dynein’s mechanical element. In contrast to the role of AAA1 in driving motility, nucleotide transitions in AAA3 gate the transmission of conformational changes between AAA1 and the linker, suggesting that AAA3 acts as a regulatory switch. Further structural and mutational studies also uncover a role for the linker in regulating the catalytic cycle of AAA1. Together, these results reveal how dynein’s two major ATP-binding sites initiate and modulate conformational changes in the motor domain during motility.

Unique double-ring structure of the peroxisomal Pex1/Pex6 ATPase complex revealed by cryo-electron microscopy

“…We therefore assume that these two sites are unoccupied and that nucleotide occupancy of the Pex1 subunits has a major effect on subunit spacing. A correlation between subunit spacing and nucleotide occupancy has also been observed in recent structures of dynein (50). In our ATPγS structure, some of the occupied sites might contain ADP, as the differences from the ADP structure are only small and ATPγS can slowly be hydrolyzed or be contaminated with ADP.…”

Cytoplasmic dynein regulates its attachment to microtubules via nucleotide state-switched mechanosensing at multiple AAA domains

“…However, recent reports relate linker conformation to dynein's ATPase activities. In the absence of load, ATP at AAA3 blocks reorientation of the linker from the post-to the prepowerstroke conformation (9). In addition, binding of the cofactor Lis1, which mechanically obstructs linker movements, uncouples AAA1's ATPase activities from changes in MT-binding affinity (44).…”

“…Although AAA3 plays an important role in controlling dynein-MT attachment (22,23,25), the details are just emerging. By probing dynein-MT interactions in the absence of load (9,34) and with force applied to the dynein C terminus (34), DeWitt et al and Bhabha et al concluded that AAA3 must be in a posthydrolysis state for ATP-induced, AAA1-mediated MT release. Our MTBR and C-terminal pulling results support these findings, but if tension is applied via the linker, then AAA1-mediated MT release is allowed when AAA3 enters the ATP state.…”

“…Finally, we dissect AAA1-and AAA3-mediated nucleotide-induced modulation of dynein's inherent response to force, identifying (i) a previously undescribed weakening of MT attachment caused by ADP binding at AAA3 and (ii) a novel function for the linker in the AAA3-mediated "gating" of the nucleotide-dependent regulation of dynein-MT binding by AAA1. When tension is absent or applied via dynein's C terminus, ATP binding to AAA1 induces MT release only if AAA3 is in the posthydrolysis state, as described recently (9,34). However, under more physiological conditions in which tension is applied to the linker, ATP binding to AAA3 is sufficient to "open" the regulatory gate.…”

“…The stalk configuration influences both MT affinity and ATPase activity (5) and thereby mediates bidirectional allosteric communication between the AAA ring and the MTBD (3,6). Finally, a ∼10-nm "linker" also emerges from the ring and undergoes cyclic reorientations that generate force and displacement (7)(8)(9).…”