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We have demonstrated a novel micro/nanotransport system using biomolecular motors driven by adenosine triphosphate (ATP). For the driving mechanism, microtubule-kinesin system, which is one of the linear biomolecular motor systems was investigated. ATP dissolved in an aqueous condition is hydrolyzed to adenosine diphosphate (ADP) to energize the bionanoactuators in this mechanism. This means the system does not require an external electrical or mechanical energy source. Therefore, a purely chemical system which is similar to the in vivo transport will be realized. This paper reports some fundamental studies to integrate biomaterials and MEMS. The microtubules, or rail molecules, were patterned on a glass substrate with poly(dimethyl siloxane) (PDMS) using a regular soft lithography technique. Microbeads (320 nm in diameter) and a micromachined structure (2×3 μm, 2 μm in thickness) coated with kinesin molecules were transported along the microtubules at an average speed of 476±56 and 308 nm/s, respectively. While ATP injection activated the transport system we have also managed to provide repetitive on/off control using hexokinase as an inhibitor. For the minimum response time in the repetitive control, the optimized concentration for ATP was 102 μM and 103 U/L for hexokinase.