Structural studies of terpenoid biosynthesis and bacterial cell division

The objective of this work is to investigate the structures of two nucleotide binding proteins: mevalonate kinase (MVK) and FtsZ. MVK is the key enzyme involved in terpenoid biosynthesis. In this study, we solved the crystal structures of the M. jannaschii MVK apoprotein, as well as the protein in complex with ligands. Its fold was analyzed and firmly established within the GHMP kinase family, in which homoserine kinase (HSK), phosphomevalonate kinase and galactokinase also belong. Structural analysis in combination with enzyme kinetics studies revealed the mechanism of this enzyme upon substrate binding, catalysis and inhibition. In particular, the phosphate-binding loop was found to be critically involved in the binding of nucleotides and terpenoids, via the interaction with a di-phosphate moiety from the ligand. An enzymatic reaction mechanism was constructed based on our structural data and it is consistent with kinetics studies from the literature. In this mechanism, the invariant residue Asp 155 functions as a general base that increases the nucleophilicity of the phosphoryl acceptor. Finally, a virtual screening study has been performed to explore the ligand binding potential of MVK. Compounds predicted to bind MVK were tested and analyzed. FtsZ is a prokaryotic homologue of tubulin that forms the apparatus for bacterial cell division. The structure of a crystal filament of the M. tuberculosis FtsZ complexed with GDP was described in this study. It shows an anti-parallel, left-handed double helical architecture. Compared with the straight crystal filament revealed earlier by other groups, the catalytic T7 loop in our structure is found to be outside the nucleotide binding site, indicating the GTPase is inactive. Furthermore, the buried surface area in our crystal filament is less, probably suggesting the helical FtsZ filament is less stable. We therefore proposed that the hydrolysis of GTP and the releasing of the ?-phosphate group will trigger the rearrangement of the FtsZ fibler, characterized by the exclusion of the T7 loop, which might lead to a less stable helical filament and would be the first step for the disassembly of FtsZ polymer.