Structural and Biochemical Studies of FrzCD, A Cytoplasmic Methyl-accepting Chemosensory Protein (MCP)

Abstract

FrzCD is a Methyl-accepting chemotaxis protein (MCP) of Myxococcus xanthus. It was recently found to colocalize with the nucleoid and aid in the cooperative response of bacteria to signals. Invitro DNA binding studies suggested the sequence-independent DNA binding is by utilizing the N-terminal basic tail. MCPs have a sensor domain which is generally periplasmic for ligand binding, a HAMP linker to amplify and transmit the signal and a signaling unit to signal the downstream pathways. Our bioinformatic analysis has found out the presence of two contiguous HAMP domains in FrzCD. We aim to investigate the role of HAMP domains by systematic designing of domain deletion constructs and performing protein oligomerization and DNA-binding studies. Our preliminary results indicate that the higher order oligomerization of protein is mediated by the coiled-coil signaling unit. In the DNA-free state, HAMP domains restrict oligomeric state of the protein to a dimer. EMSA shows that coiled-coil domain stabilizes the protein-DNA complex possibly through a higher-order array formation. We are progressing further to quantify the binding affinities, to find the oligomeric state of the protein in the DNA-bound form and to obtain the crystal structure of the protein.