Cytoplasmic dynein drives minus-end-directed motility along microtubules by converting ATP hydrolysis into coordinated structural changes but how this process produces directional stepping remains unresolved. The dynamics of dynein stepping have previously been characterized by tracking flexible regions of the motor with limited resolution. Here we site-specifically labeled the microtubule-binding domain of yeast dynein using a cysteine-light mutant and tracked stepping with submillisecond, nanometer precision at physiological ATP using minimal fluorescence photon flux (MINFLUX). We show that dynein hydrolyzes one ATP per step and moves in multiples of 8 nm. Steps are preceded by a transient plus-end-directed displacement ('dip'), corresponding to microtubule release upon ATP binding and diffusion of the stepping head around its partner. A slow ATP-hydrolyzing mutant shows more frequent dips, supporting a model in which ATP hydrolysis produces net forward movement. These results clarify the sequence of mechanochemical events underlying productive dynein stepping.
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| Evidence ID | Analyze ID | Gene/Complex | Systematic Name/Complex Accession | Qualifier | Gene Ontology Term ID | Gene Ontology Term | Aspect | Annotation Extension | Evidence | Method | Source | Assigned On | Reference |
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| Evidence ID | Analyze ID | Gene | Gene Systematic Name | Phenotype | Experiment Type | Experiment Type Category | Mutant Information | Strain Background | Chemical | Details | Reference |
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| Evidence ID | Analyze ID | Gene | Gene Systematic Name | Disease Ontology Term | Disease Ontology Term ID | Qualifier | Evidence | Method | Source | Assigned On | Reference |
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| Evidence ID | Analyze ID | Regulator | Regulator Systematic Name | Target | Target Systematic Name | Direction | Regulation of | Happens During | Regulator Type | Direction | Regulation Of | Happens During | Method | Evidence | Strain Background | Reference |
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| Site | Modification | Modifier | Source | Reference |
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| Evidence ID | Analyze ID | Interactor | Interactor Systematic Name | Interactor | Interactor Systematic Name | Allele | Assay | Annotation | Action | Phenotype | SGA score | P-value | Source | Reference | Note |
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| Evidence ID | Analyze ID | Interactor | Interactor Systematic Name | Interactor | Interactor Systematic Name | Assay | Annotation | Action | Modification | Source | Reference | Note |
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| Complement ID | Locus ID | Gene | Species | Gene ID | Strain background | Direction | Details | Source | Reference |
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| Evidence ID | Analyze ID | Dataset | Description | Keywords | Number of Conditions | Reference |
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