Molecular observations of synaptic proteins provide a unique viewpoint on neurotransmission that complements electrophysiology measurements. We developed molecular tweezers that can monitor the zippering of a single SNARE complex, a process that drives synaptic vesicle fusion. The method simultaneously manipulates and probes the structure of a SNARE complex with a millisecond time resolution, revealing partially zippered SNAREs and their interactions with other presynaptic proteins. In this way, a force application technique can be employed to address mechanical aspects of molecular neuroscience.