The catalytic mechanisms of complex biological machines may involve a combination of chemical steps and conformational transitions. The latter are oftentimes hidden to traditional biochemical investigations, but can be exposed by single-molecule experiments. Single-molecule FRET (smFRET) is an ideal tool to probe the conformational dynamics accompanying functional dynamics of biological machines (1). We recently discovered that functional motions can be very fast, occurring on the microsecond time scale. Our first experiment involved the domain closure reaction of the enzyme adenylate kinase from *E. coli *(2). Surprisingly, we found that the bound enzyme opens and closes its domains much faster than the unbound enzyme, and two orders of magnitudes faster than the turnover rate of the enzyme! This exciting finding, which radically deviates from previous observations on adenylate kinase, led us to suggest that multi-substrate enzymes use numerous cycles of conformational rearrangement as a means to optimize the mutual orientation of their substrates for reaction. We also studied the functional dynamics of the disaggregation machine ClpB from *T. Thermophilus*. This machine is comprised of six identical subunits arranged as a barrel. A coiled-coil domain resides on the outside surface of each subunit, and this domain has been implicated as the switch of the machine, to which the co-chaperone DnaK binds. We found that the coiled-coil domain resides in two conformational states, which interchange on the microsecond time scale, making it a continuous, tunable switch. Further, a complex network of allosteric interactions involving the coiled-coil domain, the protein substrate-binding site and the ATP-binding sites was revealed.