CRYSTALLOGRAPHIC ANALYSIS OF THE MAP KINASE P38. Zhulun Wang¹, Jiahuai Han² & Elizabeth J. Goldsmith^1, Department of Biochemistry, UT Southwestern Medical Center at Dallas, Dallas, TX 75235^1, Department of Immunology, The Scripps Research Institute, La Jolla, CA 92037²

The mitogen-activated protein (MAP) kinase cascades are major signaling pathways that transmit extracellular information from the cell surface into the intracellular responses. Various extracellular stimuli, such as, growth factors, heat, UV-irradiation, inflammatory cytokines and hyperosmolarity, activate MAP kineses by dual phosphorylation on threonine(T) and tyrosine(Y). Three distinct MAP kineses signal transduction pathways have been defined so far in mammalian cells based on their differential activation selectivity and substrate specificity. As a new member of the MAP kinase family, p38 has been identified in a stress-activated signal transduction pathway. The three dimensional structure of p38 and comparison of p38 to ERK2 will help to elucidate the mechanisms of specificity determination in this family.

His6-tagged recombinant murine p38(45kDa) protein has been expressed, purlfied and crystallized by vapor diffusion method in hanging-drop with 20% PEG8000 as precipitant. The typical crystal reaches 0.6 x 0.3 x 0.06mm in two weeks. The crystals belong to the orthorhombic space group P2₁2₁2₁ with unit cell dimensions of a = 46.1Å, b=86.9 Å and c=126.5 Å. Native datasets, with diffraction limit of 2.2 Å, at both room temperature and low temperature, have been collected on a Rigaku it-axis. Molecular replacement using the MAP kinase ERK2 as a model structure and multiple isomorphous replacement have been applied to solve the structure.