This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Matrix metalloproteinases (MMPs) comprise a family of zinc-dependent proteolytic enzymes that degrade and remodel all the major components of the extracellular matrix, the space between the cells. Playing such a broad physiological role the MMPs have been implicated in a host of diseases including cancer, arthritis, and cardiovascular disease. However, over the last two decades much effort and little reward have been realized toward their control. The catalytic site in all MMPs consists of a zinc ion coordinated to side chain nitrogen atoms of 3 histidine amino acids(residues). The fourth coordination site is occupied by a water molecule in the active form of the enzyme producing the [(His)3Zn(II)-L] (L=OH2) catalytic center. This water molecule replaces a cysteine sulfur atom, which is bound in the inactive enzyme, or is replaced by the zinc binding group of an inhibitor, a molecule that stops catalysis. Known critical proton transfer steps occur in the MMP mechanism of catalytic action and activation steps, and are also thought to occur during inhibition. This proposal involves theoretical calculations of the MMP active site, with the hypothesis that structural changes about the zinc ion have a significant impact on the energetics of these proton transfer processes, as outlined: To examine the geometric influences on protonation state of the MMPs active site, hybrid quantum mechanical/semi-empirical calculations will be performed on large MMP active site models. The model will encompass the entire HExGHxxGxxH sequence of conserved residues of the catalytic domain. The HExGHxxGxxH sequence is the signature zinc-binding motif of all MMPs, with the catalytic zinc ion coordinated to these 3 histidine (H) residues. The zinc ion, the three coordinated histidines, the glutamate (E in the sequence), and the fourth zinc coordinated ligand (L = OH2 or SHCH3), will be treated using density functional theory or MP2 theory, while the rest of the sequence is treated using the semi-empirical PM3 method. Zinc geometry influences on the protonation state of the zinc bound ligands will be analyzed, and key interactions will be identified detailing the molecular basis for the zinc geometry versus protonation state relationship. The hybrid calculations will utilize the ONIOM technique employed in the Gaussian 03 software and the computations will be carried out by myself and 1 or 2 undergraduate researchers.
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