This study describes the identification of the protein target of the first chemical inhibitor (RIP-1) of t he 19S regulatory particle (RP) of the 25S proteasome. Periodate-triggered chemical cross-linking of DOPA-conjugate RIP-1 and the 26S proteasome identified Sug2/Rpt4, one of the six ATPases in the 19S as the molecular target of RIP-1 for Sug2/Rpt4 was domonstarated by examining cross-linking reactions with each ATPase if tge 19S RP. RIP-1 should provide a useful biological tool to probe the various biological roles of Sug2/Rpt4.