Assembly, structure, and function of the 26S proteasome

Abstract

The 26S proteasome is a large multiprotein complex involved in the regulated degradation of ubiquitinated proteins in the cell. The 26S proteasome has been shown to control an increasing number of essential biochemical mechanisms of the cellular lifecycle including DNA synthesis, repair, transcription, translation, and cell signal transduction. Concurrently, it is increasingly seen that malfunction of the ubiquitin proteasome system contributes to the pathogenesis of disease. The recent identification of four molecular chaperones, in addition to five previously identified chaperones, have provided mechanistic insight into how this cellular megastructure is assembled in the cell. These data, together with new insights into the structure and function of the proteasome, provide a much better understanding of this complex protease.

Figure 1

Composition of the 26S proteasome. Proteasome is formed by two regulatory particles abutting cylindrically shaped core particle. Core particle is formed by two α-rings and two β-rings. Regulatory particle consists of a base complex and a lid complex. Rpn10 is at the interface between these complexes (shown in light blue). The lid (green) contains contains indicated subunits and its function is not well understood, with the exception of rpn11, which functions as a de-ubiquitinating enzyme. The base contains a ring formed by six AAA-ATPases, Rpt1–6 (purple ring), and the subunits Rpn1, Rpn2 and Rpn13 (shown as orange box; see text for more details). The proteasome contains three heteromeric ring structures, each present twice. The α and β-ring are formed by seven subunits; scissors indicate the active sites and the dots in the a ring show the binding pockets for the Rpt tails (see text). The pocket between α7 and α1 lacks a Lys typically found in the pocket and might not harbour an Rpt C-terminus. The Rpt-ring is formed by six AAA-ATPases.

Figure 2

(A) Domain topology of Nas6, Rpn14, Hsm3 and Nas2. Below the topology is shown a 3D-structure of the domains to illustrate the structural difference. Only for Nas6 a structure been determined, for the others the structure of a similar domain from a different protein have been used (pdb coordinates used 1IXV for Nas6, 2H12 for WD40 repeat, 3GRL for Armadillo like repeats, 1G9O for PDZ domain). Hsm3 shows little conservation and detailed analysis of the domain can be found in Le Tallec et al.. Surprisingly, while structurally unrelated, these chaperones bind to the same small C-domain in the AAA-ATPases. (B) The domain topology of the proteasomal AAA-ATPases. cc is coiled coil region, OB is the OB-domain, ATPase is the ATPase domain containing the walker A and walker B motifs and the C indicates the C-domain, typically found in AAA-ATPases behind the ATPase domain. The structure shown is a ring formed by the ATPase (Blue) and C-domains (red) of six proteasome-activating nucleotidase (PAN) AAA-ATPases from the archaea Methanocaldococcus jannaschii. The CP cartoon is shown to illustrate the expected interface of the CP and the ATPase ring. Cartoons and structures are not drawn to scale.

Figure 2

(A) Domain topology of Nas6, Rpn14, Hsm3 and Nas2. Below the topology is shown a 3D-structure of the domains to illustrate the structural difference. Only for Nas6 a structure been determined, for the others the structure of a similar domain from a different protein have been used (pdb coordinates used 1IXV for Nas6, 2H12 for WD40 repeat, 3GRL for Armadillo like repeats, 1G9O for PDZ domain). Hsm3 shows little conservation and detailed analysis of the domain can be found in Le Tallec et al.. Surprisingly, while structurally unrelated, these chaperones bind to the same small C-domain in the AAA-ATPases. (B) The domain topology of the proteasomal AAA-ATPases. cc is coiled coil region, OB is the OB-domain, ATPase is the ATPase domain containing the walker A and walker B motifs and the C indicates the C-domain, typically found in AAA-ATPases behind the ATPase domain. The structure shown is a ring formed by the ATPase (Blue) and C-domains (red) of six proteasome-activating nucleotidase (PAN) AAA-ATPases from the archaea Methanocaldococcus jannaschii. The CP cartoon is shown to illustrate the expected interface of the CP and the ATPase ring. Cartoons and structures are not drawn to scale.

Figure 3

Cartoon showing the components known to be involved in the RP-assembly. The pathways and order of events are still unclear. Therefore an interaction map is displayed instead. The known potential interactions between the different complexes are shown in green. The chaperones are known to interfere with the interactions indicated in red, suggesting these proteins have a quality control role in assembly. Nas2 has also been shown to stabilize Rpt4 and 5 (purple arrow). In the middle of the cartoon are the seven pockets of the CP surrounding the gate (which is shown in an open conformation, although generally this is closed without activators bound to the CP). The dark pocket indicates the only pocket without a conserved positive charge expected not to be able to host a tail. Rpt tails that dock in the pockets are shown as dark orange extensions from the Rpt proteins. Numbers 1 to 6 indicate the different Rpt proteins, H3 Hsm3, R14 rpn14, N6 Nas6 and N2 Nas2.