9.B.1 The Integral Membrane CAAX Protease (CAAX Protease) Family
Posttranslational lipidation modulates the functions of some proteins. Isoprenoids (i.e., farnesyl or geranylgeranyl groups) are attached to cysteine residues in proteins containing C-terminal CAAX sequence motifs (where A is an aliphatic residue and X is any residue). Isoprenylation is followed by cleavage of the AAX amino acid residues, and in some cases, by additional proteolytic cuts. Pryor et al. (2013) determined the crystal structure of the CAAX protease Ste24p, a zinc metalloprotease catalyzing two proteolytic steps in the maturation of yeast mating pheromone a-factor. The Ste24p core structure is a ring of seven transmembrane helices enclosing a voluminous cavity containing the active site and substrate-binding groove. The cavity is accessible to the external milieu by means of gaps between splayed transmembrane helices. They hypothesized that cleavage proceeds by means of a processive mechanism of substrate insertion, translocation, and ejection.
Isoprenoid groups are conjugated to proteins via cysteine residues of CaaX acceptor sequences in which the cysteine attachment site is followed by two aliphatic amino acid residues and one unspecified residue at the protein C-terminus. Isoprenylation is generally accompanied by two subsequent processing steps, proteolytic cleavage of the aaX residues and carboxymethylation of the newly exposed carbonyl group of the modified cysteine residue . Some isoprenylated proteins also undergo additional proteolytic processing, including an additional cleavage by the same protease that initially removes the aaX residues. At least two classes of enzymes are responsible for the cleavage of isoprenylated proteins and peptides. One of these is the ras-converting enzyme (Rce) family of Type II prenyl proteases, responsible for proteolytic processing of signal-transducing proteins including Ras and the Gγ subunits of heterotrimeric G protein complexes . The other is the Ste24p family of Type I prenyl proteases, first identified in yeast based on its role in maturation of the mating pheromone a-factor. Extensive characterization of the role of Ste24p in a-factor processing has been conducted in the yeast system. The proteolytic activity of Ste24p requires zinc, consistent with the fact that Ste24p contains the zinc metalloprotease signature motif HEXXH.
A human ortholog of Ste24p, ZMPSTE24 (Zinc MetalloProtease STE24), can complement the full function of yeast Ste24p. A substrate for ZMPSTE24 is prelamin A, the precursor to the nuclear intermediate filament protein lamin A. Lamins provide mechanical stability to the nuclear envelope, function as scaffolds for localization of other proteins and for cytoskeletal attachment, regulate chromatin, and are implicated in transcription and DNA repair and replication. Mutations in either ZMPSTE24 or the processing site of prelamin A are associated with a spectrum of premature-aging diseases referred to as progeria. The severity of different forms of progeria is reported to be correlated with extent of loss of ZMPSTE24 activity. Ste24p is localized to the endoplasmic reticulum membrane. Its proteolytic activity requires zinc, consistent with the fact that Ste24p contains the zinc metalloprotease signature motif HEXXH.
Ste24 enzymes, a family of eukaryotic integral membrane proteins, target prenylated substrates, but also some nonprenylated substrates. Reduced activity of the human ortholog, HsSte24, is linked to multiple disease states (laminopathies), including progerias and lipid disorders. Ste24 possesses a unique 'alpha-barrel' structure consisting of seven transmembrane (TM) alpha-helices encircling a large intramembranous cavity (~14 000 A(3) ). The catalytic zinc, coordinated via a HExxH...E/H motif characteristic of gluzincin ZMPs, is positioned at one of the cavity's bases. The interrelationship between Ste24 as a gluzincin, a long-studied class of soluble ZMPs, and as a novel cavity-containing integral membrane protein protease has been explored. Goblirsch et al. 2019 developed a model of Ste24 that provides a conceptual framework for this enzyme family. The model consists of an interfacial, zinc-containing 'ZMP Core' module surrounded by a 'ZMP Accessory' module, both capped by a TM helical 'alpha-barrel' module. Multiple sequence alignment of 58 Ste24 orthologs revealed 38 conserved residues, apportioned unequally among the ZMP Core, ZMP Accessory, and alpha-barrel modules. This tripartite architecture representation of Ste24 provides a unified image of this enzyme family (Goblirsch et al. 2019). Yeast Ste24 has TC# 9.B.1.1.3 while human Ste24 has TC# 9.B.1.1.3. See these TC#s and Goblirsch and Wiener 2020 for more details.