Fig. 4.

WNG1 has adapted its active site to catalyze phosphoryl transfer without a Gly-loop. (A) Sequence logos of the WNG kinase VAIK, HRD, and DFG motifs indicate conservation of critical catalytic residues. (B) A homology model of the WNG1 structure based on the BPK1 crystal structure, (gray and blue) has been superimposed with the structure of PKA (1.96-Å backbone rmsd; 529 atoms compared). For clarity, only the PKA Gly-loop (orange) and bound nucleotide are shown. (C) A model of the WNG1 active site structure, colored as in B. Bound ATP has been modeled based on superposition of the PKA structure. Residues that comprise either canonical motifs or WNG-specific substitutions are annotated and shown as sticks. (D) Kinase activities of WT WNG1 and the indicated mutant proteins using MBP as a protein substrate, quantified by ³²P scintillation. Motifs altered by the mutants are shown above the data points. (E) A representative Michaelis–Menten fit of in vitro kinase assays of WNG1 using MBP as a substrate while varying ATP concentration.