A significant challenge in utilizing kinesin biomolecular motors in integrated nanoscale systems is the ability to regulate motor function in vitro. Here we report a versatile mechanism for reversibly controlling the function of kinesin biomolecular motors independent of the fuel supply (ATP). Our approach relied on inhibiting conformational changes in the neck-linker region of kinesin, a process necessary for microtubule transport. We introduced a chemical switch into the neck-linker of kinesin by genetically engineering three histidine residues to create a Zn2+-binding site. Gliding motility of microtubules by the 2 mutant kinesin was successfully inhibited by >= 10 mu M Zn2+, as well as other divalent metals. Motility was successfully fully restored by removal of Zn2+ using a number of different chelators. Lastly, we demonstrated the robust and cyclic nature of the switch using sequential Zn2+/chelator additions. Overall, this approach to controlling motor function is highly advantageous as it enables control of individual classes of biomolecular motors while maintaining a consistent level of fuel For all motors in a given system or device.