Structural Analysis of the MAP Kinase ERK2 and Studies of MAP Kinase Regulatory Pathways

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Publisher Summary This chapter primarily highlights structure–function studies performed collaboratively between the Goldsmith and Cobb laboratories at the University of Texas Southwestern Medical Center in Dallas and includes experiments examining the relationship of MEKKl to the MAP kinase pathway and potential feedback mechanisms in the pathway. Mechanisms regulating the MAP kinase pathway are complicated and inactidvation methods plentiful. Receptor tyrosine kinases regulate the pathway through Ras; heterotrimeric G protein-coupled receptors also use Ras to activate the pathway, although there may also be Ras-independent mechanisms. The MAP kinase pathway is stimulated by numerous hormones, growth factors, and oncogene products including Ras and contributes to their spectrum of actions. The MAP kinases, however, are pleiotropic, phosphorylating many substrates throughout the cell. This pathway has been repeated several times in yeast and mammalian cells, although mechanisms regulating the similar but parallel cascades are sketchier. There are three low-activity forms of each enzyme, the unphosphorylated protein and the two singly phosphorylated forms, that contain phosphate on only tyrosine or only threonine. These two singly phosphorylated ERKs have little more protein kinase activity than unphosphorylated proteins. Because ERKl and ERK2 can autophosphorylate on tyrosine, the form containing only threonine phosphate may, in as yet undefined circumstances, be able to reactivate itself through autophosphorylation. The three-dimensional structure of ERK2 contains the two-domain organization found in all protein kinases whose structures have been determined thus far. The smaller N-terminal domain provides many contacts for ATP, and the larger C-terminal domain contains the major determinants for protein substrate interactions. It is very important to identify the unique features, if any, of these enzyme pairs. It is interesting that phosphorylation by MAP kinase had so little effect on MEK activity, because phosphorylation of MEKl by cdc2 at a site (S286) close to the MAP kinase site (T292) in the C-terminal insert greatly inhibits MEKl activity.