Interior modification through preferential substitution of heteroatoms in 2D materials can rescue surface reactive sites for enhanced charge kinetics and photo-activity. Here, Ru and Ni heteroatoms were successfully incorporated in ZnIn2S4 (ZIS) nanosheets via a simple hydrothermal technique. Ru-ZIS photocatalysts exhibited improved photo–water splitting activity (26,400 µmol. h−1g−1) and good stability than that of Ni-ZIS and pristine ZIS under simulated sunlight, which may be related to the increase in reactive sites with Ru substitution, optimized adsorption-free energy of the reaction intermediates and charge kinetics. Ru substitution influenced Zn site availability and thus coordinated bonding with surrounding S ions. Ru-ZIS showed excellent hydrogen evolution performance and minimal change in Gibbs free energy (ΔGH, ∼0 eV). By analyzing the density of states, both Ru and S sites had greater H* intermediate absorption abilities. This study elaborates on the effect of preferential heteroatom substitution on photocatalytic performance and has widespread applications.