The typical function of kinesins is to transport cargo along microtubules. Binding of ATP to microtubule-attached motile kinesins leads to cargo displacement. To better understand the nature of the conformational changes that lead to the power stroke that moves a kinesin's load along a microtubule, we determined the X-ray structure of human kinesin-1 bound to αβ-tubulin. The structure defines the mechanism of microtubule-stimulated ATP hydrolysis, which releases the kinesin motor domain from microtubules. It also reveals the structural linkages that connect the ATP nucleotide to the kinesin neck linker, a 15-amino acid segment C terminal to the catalytic core of the motor domain, to result in the power stroke. ATP binding to the microtubule-bound kinesin favors neck-linker docking. This biases the attachment of kinesin's second head in the direction of the movement, thus initiating each of the steps taken.