PRODUCTION AND CHARACTERIZATION OF NOVEL INTRINSICALLY ACTIVE VARIANTS OF THE YEAST HOG1 AND THE MAMMALIAN p38? MAP KINASES

Abstract

Mitogen-activating protein kinases (MAPKs) play critical roles in transducing a variety of signals. It is difficult to attribute any output to a given MAPK because several MAPKs and other pathways are co-activated. Activation of a given MAPK individually would disclose the function of this MAPK. This could be achieved by expression of intrinsically active variants of MAPKs. The goal is to develop such variants of the yeast Hog1 and the human p38gamma MAPKs and study their mechanism of action. Previous studies showed that particular mutations in a structural element termed the ?hydrophobic core? rendered Hog1/p38 intrinsically active. I introduced more point mutations to this core. Each mutation rendered Hog1 intrinsically active suggesting that the core is an inherent structural locker of spontaneous activity. I introduced 20 point mutations to p38gamma and characterized them in vitro and in cell culture. None of the mutations seemed to render the kinase intrinsically active.