The viscera of narrow-barred Spanish mackerel (Scomberomorus commerson) fish contains abundant alkaline and acidic proteases that can be isolated using various methods. We purified pepsinogen from S. commerson viscera using ammonium sulfate precipitation, Sephadex G-50 gel filtration, and DEAE-cellulose ion-exchange chromatography. Two peaks corresponding to pepsinogen were observed during gel filtration and further purified using ion-exchange chromatography. Additionally, we investigated the effects of polyethylene glycol graphene oxide (GO-PEG) at a concentration of 3 mg/mL on enzyme activity and stability. Results demonstrated a significant enhancement in enzyme activity and increased stability at higher temperatures compared to the control group. These findings suggest that GO-PEG can improve enzyme production and stability. However, purified pepsin was found to be inactive in the presence of 0.1 M pepstatin A. In conclusion, S. commerson viscera offers a valuable source for pepsin purification. The purification process involving ammonium sulfate precipitation, gel filtration, and ion-exchange chromatography successfully obtained purified pepsinogen. The use of GO-PEG exhibited promising potential in enhancing enzyme activity and stability, highlighting the role of nanosheets in improving enzyme thermal stability.