Cytoplasmic dynein is a microtubule (MT) motor protein comprising two classes: dynein-1 and dynein-2. We purified recombinant human dynein-1 and dynein-2 from HEK-293 cells by expressing the streptavidin-binding peptide-tagged human cytoplasmic dynein-1 and dynein-2 heavy chains (HCs), respectively. Electron microscopy of the purified molecules revealed a two-headed structure composed of characteristic dynein motor domains. In an in vitro MT gliding assay, both dynein-1 and dynein-2 showed minus-end-directed motor activities. This is the first demonstration of dynein-2 motor activity, which supports the retrograde intraflagellar transport role of dynein-2. Our expression system of dynein HCs provides a useful m... More