Synthetic protein switches are proteins that can be controlled by an external input, and are useful tools to probe protein function. Antibody fragments and antibody mimetics can be selected to bind, activate, or inhibit several diverse protein targets. Nevertheless, their affinity for the target is often high and the binding non-reversible. Turning antibody fragments and antibody mimetics into synthetic protein switches would provide a useful tool for controlling their affinity with external cues, such as small molecules or light. This thesis explores several methods of controlling protein function. The affinity of antibody fragments and antibody mimetics for their targets could be controlled by fu-sion with a circularly permutated dihydrofolate reductase (cpDHFR). This was used to develop ligand-based affinity modulators of antibody fragments ("LAMAs"), and reversibly release and recruit GFP-fused proteins to different anchored locations in live cells upon the addition of a small molecule. Other attempts to controlling protein function are also described: (i) a covalent steric block was used to block an antibody fragments binding site for GFP on the addition of a small molecule, (ii) and a strate-gy for controlling GFP-fused receptors by delivering photoactivatable small molecules with antibody fragments was also developed. Together, these synthetic protein switches provide tools to probe several biological functions with different spatiotemporal resolutions.