Effect of C-terminal truncations on MLK7 catalytic activity and JNK activation

Mixed Lineage Kinase 7 (MLK7) is a MAPKKK with enriched expression in heart and skeletal muscle that functions to activate JNK and p38. The MLKs have several conserved domains, including a leucine zipper that in other family members mediates oligomerization critical for catalytic activity and JNK activation. Nested C-terminal deletion mutants of MLK7 from 436 to 286 as well as a mutant lacking only the leucine zipper (delLZ) were generated to determine the role of these domains in catalytic activity and JNK activation. Specific activity of MLK7366 was 75% full length while 436, 322, and delLZ retained approximately 25% and 286, 4% of the full-length catalytic function, demonstrating that the leucine zipper, while not absolutely necessary for catalytic activity, is required to reach full catalytic function of the Enzyme. Co-transfection studies of JNK with the MLK7 mutants demonstrated full JNK activation with MLK7, 436, and delLZ, marginal activation for 1-400 or 1-366, and no activation for 1-322, demonstrating that the leucine zipper is not required for JNK activation and that sequence contained in C-terminal residue 322-436 is necessary for full pathway activation by MLK7.