W0443

Time-resolved Crystallographic Studies of the Heme-based Sensor Protein FixL. Jason Key¹, Vukica Srajer^1,2, Reinhard Pahl², and Keith Moffat^1,2, ¹Dept. of Biochemistry and Molecular Biology, The Univ. of Chicago, 920 E. 58^th St., Chicago, IL 60637 & ²Consortium for Advanced Radiation Sources, The Univ. of Chicago, 5640 S. Ellis Ave, Chicago, IL 60637.

The protein FixL, a heme-based oxygen sensor found in nitrogen-fixing Rhizobia, is responsible for the regulation of nitrogen fixation genes in response to molecular oxygen. It is a modular protein composed of two domains: an N-terminal PAS domain which contains covalently bound heme (FixLH), and a C-terminal histidine kinase enzymatic domain whose activity is regulated by the N-terminal sensor domain in response to heme-bound ligand. The sensory domain of FixL is a member the emerging family of heme-based PAS domain gas sensor proteins, distinct from globins in structure and function. In order to investigate the early structural events of signal transduction in FixLH, we are conducting time-resolved crystallographic experiments on the photolabile CO complex of FixLH from the organism Bradyrhizobium japonicum using time-resolved Laue diffraction. Structures of the FixL heme domain have been determined at room temperature to investigate structural changes upon CO binding as well as to provide phase information for room temperature time-resolved studies. These time-resolved experiments reveal structural details of ligand discrimination in the hydrophobic FixLH binding pocket during CO recombination, and may reveal possible docking sites or ligand transit pathways in this important class of molecules.