Protein levels in cells can be controlled by delivering and releasing the target protein to a cell. Herein, we report the preparation of photoresponsive nanoparticles with different protein loadings for application as protein delivery vehicles. The nanoparticles were prepared by radical polymerization using an X-shaped photoresponsive linker (Mw = 2000) and a cationic linker. The incorporation of a fluorescent polysaccharide (FITC-Dex) or protein (caspase 3) into the nanoparticles did not create significant differences in the physical properties of the resultant nanoparticles; they were ~150 nm in size with weak positive zeta potentials. These new photoresponsive nanoparticles can encapsulate and release various protein-containing compounds with different molecular weights (3–250 kDa). In addition, the amount of compound released from the nanoparticles can be tailored by altering the amount of compound encapsulated in the nanoparticles. In this study, nanoparticles containing cationic units were introduced into cells via endocytosis. We controlled the activity of the nanoparticle-released proteins within HeLa cells. Specifically, adjusting the protein loading in the nanoparticles enabled control over the activity of the proteins released into the cell, and thus the level of protein function, which provides a powerful tool for elucidating the functions of proteins.