Stereocilia are mechanosensitive organelles projecting from the surface of inner ear hair cells that detect nanometer deflections induced by sound, gravity or head movement. Unconventional myosin-15 (encoded by Myo15) is hypothesized to regulate stereocilia development by delivering cargoes such as whirlin and Eps8 to their tips. Whether myosin-15 actually functions as a transporter remains untested and little is known regarding its activity, structure and regulation within the highly specialized stereocilia compartment.We have characterized the biochemical kinetics of a subfragment-1 (S1) like truncation of mouse myosin-15, comprising the catalytic ATPase/actin binding domain plus two IQ light chain binding sites. Expression of the recombinant S1 fragment in Sf9 insect cells required co-expression of multiple molecular chaperones in order to recover significant quantities of active protein. Unlike most unconventional myosin classes, the IQ regions of myosin-15 did not bind calmodulin with high affinity; instead preferentially associating with essential (MYL6) and regulatory (MYL12A) light chains that typically partner with myosin-2 isoforms. Single molecule TEM confirmed that the purified S1 was correctly folded and monomeric. The S1 fragment was mechanically active and moved actin filaments at ∼170 nm·s−1 in a gliding motility assay. A powerstroke displacement of 7.8 nm was measured by optical trapping. Transient kinetics revealed that ATP binding to myosin-15 was slow and weak (K1k2 = 0.17 μM−1s−1, 1/K1 = 1898 μM), and that ADP release (∼12s−1) was the rate-limiting step; indicating the predominant steady-state intermediates for myosin-15 would either be with ADP, or no nucleotide bound. Both states bind strongly to actin, suggesting that myosin-15 could be capable of longer-range processive motility if oligomerized in vivo. These data enable future kinetic and structural studies to investigate how deafness-associated mutations targeted within the ATPase ultimately disrupt stereocilia function.