Extended Data Figure 10. Expanded model of the complete cycle of substrate processing by the human 26S proteasome.

The cartoon summarizes the concept of three principle modes of coordinated ATP hydrolysis observed in the seven states and our proposal of how they regulate the complete cycle of substrate processing by the proteasome holoenzyme. Coordinated ATP hydrolysis in modes 1, 2 and 3 features hydrolytic events in two oppositely positioned ATPases^8,18, in two consecutive ATPases^6,19,52, and in only one ATPase at a time^{20–23,53–56}, respectively. Substrate processing undergoes three major steps before CP gate opening for processive translocation: (1) ubiquitin recognition; (2) simultaneous deubiquitylation and substrate engagement with the AAA-ATPase ring; and (3) translocation initiation, which involves multiple simultaneous events, including ubiquitin release, ATPase repositioning and switching of Rpt C-tail insertion pattern. In some cases, the initiation of translocation may precede deubiquitylation. In steps 1 and 2, the ATPases follow the mode-1 ATP hydrolysis. In step 3, they follow the mode-2 ATP hydrolysis. After the gate is open, the AAA-ATPases hydrolyze ATP in mode 3, in which only one nucleotide is hydrolyzed at a time.