Autophagy, a conserved “self-eating” process for cellular homeostasis, plays a crucial role in the hepatitis B virus (HBV) life cycle, including viral assembly, envelopment, release, and degradation. Autophagy can also be hijacked by HBV for its persistence and survival. However, the association between HBV mutations and autophagy remains unclear. Herein, we found that an HBV surface antigen (HBsAg) mutation involving sL13H substitution impaired HBsAg expression and caused its abnormal distribution in vitro and in a hydrodynamic injection (HI)-based mouse model for acute HBV infection. The sL13H mutation also decreased autophagosome formation and inhibited autophagic flux. Further mechanistic analysis revealed that sL13H suppresses HBV-induced autophagy initiation by inhibiting the eukaryotic translation initiation factor 2 alpha kinase 3 (EIF2AK3/PERK)-eukaryotic translation initiation factor 2 subunit alpha (EIF2S1/eIF2α)-autophagy related (ATG)5/12 axis. Therefore, our findings reveal a potential role for HBV mutations in manipulating host autophagic flux for virus persistence and pathogenesis in chronic hepatitis B.