PROSITE documentation PDOC52101

Polyphosphate kinase family-2 (PPK2) domain profile

Polyphosphate (polyP) is an inorganic linear polymer of tens to thousands of phosphoryl monomers and is found in all branches of life. The polymer is linked by high energy phosphoanhydride bonds (P-O-P). These energy-rich bonds render polyP a common polymer for energy and phosphate storage in various organisms. The biological functions of polyP are numerous, including energy storage, metal chelation, buffering, DNA uptake, gene regulation, and the bacterial stringent response to nutrient deficiency. In nature, polyP is synthesized and degraded by polyphosphate kinases (PPKs) that catalyze the reversible transfer of the terminal phosphoryl residue from nucleoside 5'-triphosphates to polyP. Two large, structurally unrelated families of bacterial PPKs have been characterized so far: PPK1 enzymes that favor polyP synthesis and the PPK2 family that favors nucleotide phosphorylation. PPK2s can be phylogenetically subdivided into three classes: class I preferentially phosphorylates nucleoside diphosphates, class II converts nucleoside monophosphates into diphosphates, and class III can phosphorylate either nucleoside mono- or diphosphates. A small subgroup of PPKs that favors the phosphorylation of pyrimidine nucleobases instead of purine nucleobases, which has been designated as PPK3, clusters phylogenetically with the PPK2 family (class I). PPK2 enzymes can be further classified into one- or two-domain enzymes, with the latter consisting of two fused PPK2 domains, probably resulting from a gene duplication event. The classes PPK2-I and -III consist of one-domain enzymes, the PPK2-II class includes both one- and two-domain proteins. The C-terminal domain of 2-domain PPK2 proteins is more conserved and shows higher similarity to the sequences of 1-domain PPK2 proteins than their N-terminal domains. The N-terminal domain of 2-domain PPK2 proteins seems to be catalytically inactive and might be responsible for the protein dimerization. Four 1-domain PPK2 proteins assemble as a D2 tetramer, whereas the 2-domain PPK2 proteins form a dimer with the four PPK2 domains arranged like a pseudotetramer [1,2,3,4,5,6].

The PPK2 domain ~230 residues in length belongs to the family of the P-loop kinases, which share a conserved globular fold with a parallel β-sheet surrounded by α-helices and the lid module (see <PDB:6B18>). P-loop kinases are characterized by the presence of two conserved sequence motifs: Walker A (GxDxxGK) involved in the binding of polyP and a Mg(2+) ion and Walker B (DRS) that coordinates the nucleotide phosphate groups. The PPK2 domain is arranged into a 3-layer α/β/α sandwich with a central 5- or 6-stranded parallel β-sheet flanked by α-helices on both sides and on the top (lid module). The potential catalytic mechanism for PPK2 enzymes involves two Mg(2+) ions. The active site is covered by the lid module, which provides several positively charged residues for the coordination of the reactive polyP chain end, as well as an Asp residue coordinating one of the Mg(2+) ions (Mg1). An internal substrate channel connects the polyP and nucleotide binding sites near the catalytic Asp (Walker A). PPK2 domains orient the terminal phosphates of both substrates (nucleotide and polyP) optimally for a nucleophilic attack of the nucleotide α- (in AMP) or β-(in ADP) phosphate oxygen on the terminal polyP phosphorus atom. This is accomplished through coordination of phosphate groups by a bridging Mg(2+) ion (Mg2). In the transition state, the transferred phosphoryl group of polyP is stabilized by the side chains of two conserved positively charged residues: an Arg from the lid module, and a Lys from the bottom of the active site, with the phosphoryl group likely remaining attached to these side chains over the entire transfer path [1,2,3,4,5,6].

The profile we developed covers the entire PPK2 domain.