Size tunable amphiphilic NPs composed of poly(gamma-PGA) and hydrophobic amino acids, such as Phe or Trp, were prepared. To prepare these size-regulated NPs, gamma-PGA-g-Phe or gamma-PGA-g-Trp dissolved in DMSO was added to various concentrations of NaCl solution. The gamma-PGA-Phe and gamma-PGA-Trp formed monodispersed NPs, and the size of NPs can be easily controlled by NaCl concentration. The different-sized NPs showed the same structure. The encapsulation of protein into the different-sized NPs was successfully achieved and the size of protein-encapsulated gamma-PGA-Phe NPs was increased when protein was encapsulated.