Fig. 2.

Pseudoenzymes can adopt multiple structural conformations, permitting them to function as molecular switches. a The pseudokinase domain of mouse MLKL was crystallised in an open (equivalent to catalytically inactive) conformation in which the activation loop adopted an unusual helix that buttresses against, and shuns, the αC helix. Counterparts of the catalytic residues in conventional protein kinases (K219, N318, E338), and the K219-interacting Q343 from the activation loop helix, are shown as yellow sticks. PDB accession 4BTF [18]. b The structure of the human MLKL pseudokinase domain crystallised in a distinct closed conformation which resembles that of an active conventional protein kinase, suggesting that MLKL has evolved to function as a catalytically inactive conformational switch. Counterparts of the catalytic residues in conventional protein kinases (K230, K331, E351) are shown as yellow sticks. The canonical αC helix glutamic acid (E250), rather than the activation loop residue observed in the mouse structure, interacts with K230, as is typical of active protein kinase structures. PDB accession 4MWI [14]. Accompanying mutational analyses illustrated that nucleotide binding by MLKL is mediated by different pseudoactive site determinants and that K219 (mouse) and K230 (human) have evolved unexpected functions to permit nucleotide binding, which might drive or inhibit a switch mechanism that controls release of the MLKL four-helix bundle (4HB) domain (shown in a) to induce cell death by necroptosis [14, 18]. Cartoons drawn using Pymol