Resonance Raman Studies of Lactoperoxidase, Its Catalytic Intermediates and Model Compounds

Lactoperoxidase is a mammalian enzyme that is part of our natural defense system. Very little is known about this enzyme, and so studies have been designed to gain an understanding of how this enzyme works in nature. Enzymes are responsible for catalyzing biochemical reactions. Often times the enzyme\u27s shape changes during its enzymatic cycle. Resonance Raman (RR) spectroscopy is capable of monitoring the structural changes occurring during this process. By examining the enzyme at various points in time one can develop a picture of how the enzyme changes as the biochemical reaction proceeds. By determining what the enzyme looks like before, during and after the reaction, one can then gain an insight into how this enzyme behaves. In some cases it is possible to construct more simple model compounds which mimic these catalytic intermediates. These studies help to characterize the enzyme in a variety of different conformations and, hopefully, the information gained here will be systematically applicable to other mammalian enzymes. All attempts to generate and monitor the enzymatic intermediates of lactoperoxidase have been unsuccessful. Currently, little is being done on RR studies of these intermediates owing to their inherent photo-lability, fast reaction times and large-scale quantities of sample required for these experiments. Recent advances made in the area of time-resolved resonance Raman spectroscopy provide an opportunity to structurally characterize the enzymatic intermediates of lactoperoxidase. The other area of interest centers on studying the factors which dictate the reactivity of the prosthetic group, i.e., heme chromophore, of the lactoperoxidase enzyme. Recently, a method for extracting the heme prosthetic group of lactoperoxidase, without modification, has been published. The inherent problem with removing the heme prosthetic group of lactoperoxidase is that the surrounding apoprotein matrix is covalently linked to the heme through the 1 and 5 methyl substituents. These covalent linkages, which occur only for the mammalian peroxidases, have hindered its removal from the protein, preventing systematic studies of its inherent reactivity