Deregulation of mitogen-activated protein kinase at low pH due to a structural rearrangement of activation segment

Abstract

Autophosphorylation of recombinant mitogen-activated protein kinase (MAPK) on Tyr was found to be several-fold stimulated at weakly acidic pH (5.5–6.0), whereas the phosphorylation of a protein substrate, myelin basic protein, was greatly inhibited at pH below 6.0. In contrast to phosphorylation at pH 8.0, both MAPK autophosphorylation and MAPK phosphorylation with upstream MAPK kinase at low pH failed to stimulate essentially its kinase activity towards the exogenous protein substrate. Immunoprecipitation and ELISA with an activation segment-specific antibody, kinetic analysis, and reversible phosphorylation assay revealed a difference in the folding of MAPK activation segment at pH 5.5 and 8.0. The data suggest that a rearrangement of the activation segment at low pH promotes a stable low-activity conformation of the enzyme which is favorable for intramolecular autophosphorylation. In this conformation, the phosphorylation of the exogenous protein substrate is inhibited due to persistent blocking of the enzyme catalytic center by the activation segment.