AAA proteins

AAA or AAA+ is an abbreviation for ATPases Associated with various cellular Activities. It is a protein superfamily in which the member proteins share a highly conserved module of approximately 230 amino acid residues.

The characteristic of AAA proteins is the coupling of chemical energy by ATPase, provided by ATP hydrolysis, to mechanical force exerted on some macromolecular substrate. This usually requires a conformational change in the AAA protein in question.

Members of the AAA family are found in all organisms and they are essential for many cellular functions.

One type of AAA Proteins are AAA Proteases, where the energy from ATP hydrolysis is used to translocate a Protein inside the Protease for degradation.

AAA-type ATPases constitute a large family of enzymes. AAA proteins are characterised by the presence of 200-250 amino-acid ATP-binding domains that contain Walker A and B motifs. AAA proteins themselves belong to the superfamily of P-loop NTPases.

Domain structure of AAA-type ATPases
AAA proteins have an N-terminal Non-ATPase domain which is followed by either one or two AAA domains (D1 and D2). In some proteins with two AAA domains, both are evolutionarily well conserved (like in Cdc48p/97). In others, either the D2 domain (like in Pex1p and Pex6p) or the D1 domain (in Sec18p/NSF) is better conserved in evolution.

From AAA to AAA+
The classical AAA family has been expanded by inclusion of a number of more distantly related cellular regulators and termed AAA+ family of ATPases (112). AAA+ proteins are involved in protein degradation, membrane fusion, DNA replication, microtubule dynamics, disassembly of protein complexes and protein aggregates.

AAAs are mechanoenzymes that manipulate the structure of substrate proteins and thereby unfold them or disassemble protein complexes.

AAAs are often Hexamers
The physiologically active form of these enzymes is often an homo-hexamer. The hexameric enzymes have an overall shape that resembles a double-ring with a central pore that might be involved in substrate processing. In the hexameric configuration, the ATP-binding site is positioned at the interface between the subunits. Upon ATP binding and hydrolysis, AAA enzymes undergo conformational changes in the AAA-domains as well as in the N-domains. These motions can be transmitted to substrate protein.

Prokaryotic AAAs
AAA proteins are not restricted to eukaryotes. Prokaryotes have AAA which combine chaperone with proteolytic activity, for example in ClpAPS complex, which mediates protein degradation and recognition in E. coli. The basic recognition of proteins by AAAs is thought to occur through unfolded domains in the substrate protein. In HslU, a bacterial ClpX/ClpY homologue of the HSP100 family of AAA+ proteins, the N- and C-terminal subdomains move towards each other when nucleotides are bound and hydrolysed. The terminal domains are most distant in the nucleotide-free state and closest in the ADP-bound state. Thereby the opening of the central cavity is affected.

AAAs in protein transport
The AAA-type ATPase Cdc48p/p97 is perhaps the best-studied AAA protein. Misfolded secretory proteins are exported from the endoplasmic reticulum (ER) and degraded by the ER-associated degradation pathway (ERAD). Nonfunctional membrane and luminal proteins are extracted from the ER and degraded in the cytosol by proteasomes. Substrate retrotranslocation and extraction is assisted by the Cdc48p(Ufd1p/Npl4p) complex on the cytosolic side of the membrane. On the cytosolic side, the substrate is ubiquitinated by ER-based E2 and E3 enzymes before degradation by the 26S proteasome.

Targeting to multivesicular bodies
Multivesicular bodies are endosomal compartments that sort ubiquitinated membrane proteins by incorporating them into vesicles. This process involves the sequential action of three multiprotein complexes, ESCRT I to III. Vps4p is a AAA-type ATPase involved in this MVB sorting pathway. It had originally been identified as a ”class E” vps (vacuolar protein sorting) mutant and was subsequently shown to catalyse the dissociation of ESCRT complexes. Vps4p is anchored via Vps46p to the endosomal membrane. Vps4p assembly is assisted by the conserved Vta1p protein, which regulates its oligomerzation status and ATPase activity.