ADP-ribosylhydrolases (ARH1, ARH2 and ARH3) are a family of enzymes to catalyze ADP-ribosylation, a reversible and covalent post-translational modification (PTM). There are four phosphorylated sites (Tyr-4, Tyr-19, Tyr-20, and Tyr-205) in ARH1. To explore the structural changes and functional impact induced by phosphorylation, molecular dynamics (MD) simulations and steered molecular dynamics (SMD) simulations were performed for the phosphorylated and non-phosphorylated ARH1 with the ligands. MD simulations results indicate that: (1) Glu-25 is more frequently in the α helix group in the phosphorylated state with the adenosine-5-diphosphate-ribosylarginine (ADP-RA) complex (51.56%) than that of the non-phosphorylated state(2.12%); (2) Ser-124 and Ser-264 become less flexible in the phosphorylated state with ADP-RA complex, which helps two residues form hydrogen bonds with ADP-RA; and (3) Tyr-211 is also less flexible in the phosphorylated state with ADP-RA complex, which helps stabilize the cation-π interaction of Y211-R119. All these changes facilitate ADP-RA to bind ARH1. In addition, according to the crystal structure of adenosine-5-diphosphate-ribose (ADP-ribose) in complex with non-phosphorylated and phosphorylated ARH1, the possible unbinding pathways of ADP-ribose from non-phosphorylated and phosphorylated ARH1 were explored respectively using SMD simulations. Our results show that phosphorylated ARH1 has more ordered structures than the non-phosphorylated type.