MCD and MLD Studies of Cytochrome Oxidase and other Hemoproteins

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9506817 Peterson This research will develop magnetic linear dichroism (MLD) as a valuable tool for the study of cytochrome c oxidase and some other metalloproteins. MLD is not sensitive to heme a or CuA(which give rise to features that dominate the magnetic circular dichroism MCD spectra of most cytochrome c oxidase derivatives one would like to study) but is sensitive to some states in which the heme a3-CUB binuclear pair can be prepared. Consequently, it is expected that the site of interest can be observed free of spectral interference from the other chromophores using the MLD approach. The major limitation associated with magneto-optical measurements on hemoproteins in general has been that the MCD signals derived from species having a low spin ferric (S = 1/2) ground-state are the most intense and can therefore dominate much of the spectrum. Such hemes tend to be simply electron transfer centers and not directly involved in substrate binding. Since for most purposes we are primarily concerned with the latter, this represents something of a problem. Nearly always, it is possible to prepare one or more interesting derivatives where the electron transfer centers are low-spin ferrous (S = O) and the substrate binding heme has some paramagnetic ground-state (S =2 usually). In these circumstances, only the active site heme has a readily measurable MCD signal. Unfortunately, there invariably remain a number of derivatives which could usefully be studied by MCD spectroscopy if it were not for small amounts of residual low-spin ferric hemes dominating the spectrum. This tends to be the rule rather than the exception and is definitely the most serious source of spectral interference associated with the technique. MLD spectroscopy represents something of a breakthrough in this regard, since it is not sensitive to either low-sin ferric, or ferrous, systems; but is likely to prove particularly useful for studying integer spin paramagnets. The MCD instrumentatio n now on-line at The University of Alabama operates between 1.2 and 300 K, O to 7 tesla, over the wavelength range 350 to 3000 nm. No other single instrument has these capabilities. Also, unlike other MCD instruments, it has been constructed entirely by American companies. The planned addition of the MLD option will represent a further unique feature in a single instrument. This equipment arguably represents something of a national resource. %%% This research will develop the technique known as magnetic linear dichroism (MLD) as a tool for the study of hemoprotein and other samples which have proved difficult to investigate by established methods. Presently, there seem to be no useful MLD data reported for any metalloprotein samples.The information obtained will assist in the elucidation of the mechanism by which certain metalloenzymes function and is, therefore, of some fundamental significance to biochemistry. The planned addition of the MLD option to the magnetic circular dichroism (MCD) instrumentation now on-line at The University of Alabama will represent a further unique feature in a single instrument and, arguably, represents something of a national resource. ***