The RING domain of MUL1 (RING MUL1 ) alone mediates ubiquitylation of the p53-transactivation domain (TAD p53 ). To elucidate the mechanism underlying the simultaneous recruitment of UBE2D2 and the substrate TAD p53 by RING MUL1 , we determined the complex structure of RING MUL1 :UBE2D2 and studied the interaction between RING MUL1 and TAD p53 in the presence of UBE2D2-UB thioester (UBE2D2~UB) mimetics. The RING MUL1 -binding induced the closed conformation of UBE2D2 S22R/C85S -UB K48R oxyester (UBE2D2 RS -UB R OE ), and strongly accelerated its hydrolysis, which was suppressed by the additional N77Amutation of UBE2D2. Interestingly, UBE2D2 S22R/N77A/C85S -UB K48R oxyester (UBE2D2 RAS -UB R OE ) already formed a closed conformation in the absence of RING MUL1 . Although TAD p53 exhibited weak binding for RING MUL1 or UBE2D2 alone, its binding affinity was enhanced and even further for RING MUL1 :UBE2D2 and RING MUL1 :UBE2D2 RAS -UB R OE , respectively. The recognition of TAD p53 by RING MUL1 as a complex with UBE2D2~UB is related to the multivalency of the binding events and underlies the ability of RING MUL1 to ubiquitylate the intrinsically disordered protein, TAD p53 .