NMR AND KINETIC STUDIES ON AZURIN
Abstract
Structural and kinetic aspects of azurin have been investigated with NMR and copper uptake kinetics. By using the nickel(II) derivative of azurin, two novel structural effects of azurin have been found. The geometry about the Ni(II) atom in Ni(II) azurin was found to be pseudotetrahedral, implying a very rigid metal site is present in azurin. The first spectroscopic evidence for a pH dependent conformational change involving histidine('35) of azurin was also found. Through studying the kinetics of copper(II) uptake by apoazurin, it was first found necessary to develop a new milder apoazurin preparation proceedure, replacing the apparently detrimental current literature preparation. It was found that the mechanism of Cu(II) uptake is much more complex than previous studies indicated. Kinetic evidence was obtained indicating that the rate determining step of the reaction changes with conditions (pH and temperature) indicating a complex mechanism. a partial mechanism explaining some of the kinetic results was computer-modeled and found to be fairly accurate. The partial mechanism proposed contradicts the previous findings, in which it has now been found that the formation of an intermediate occurs but this is not necessarily a fast process.
Degree
Ph.D.
Subject Area
Chemistry
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