pH is a critical parameter in biological systems, with acidic environments often serving as hallmarks of pathological conditions such as cancer, infection, and metabolic disorders. Here, we developed a pH-responsive synthetic cell capable of protein synthesis and release under acidic conditions. The system was constructed by integrating three molecular modules: a proton channel for pH sensing, a pH-responsive single-stranded DNA (ssDNA) that releases a trigger ssDNA upon acidification, and a toehold switch RNA that activates translation in response to the trigger ssDNA. During integration, we discovered that adjusting the annealing length between the pH-responsive and trigger strands was critical for enabling the acid-triggered protein synthesis. Using this strategy, we successfully demonstrated acid-responsive protein expression within synthetic cells. To further explore applications, we embedded the synthetic cells in a hydrogel to endow pH-responsive behavior to materials and coupled pH-responsive protein translation with a cell-penetrating peptide technology for selective release of proteins.