[16] Circular dichroism spectroscopy of hemoproteins

This chapter presents background for interpreting the circular dichroism (CD) spectra of hemoproteins, with emphasis on the CD spectrum of heme, some aspects of methodology, and several specific applications. It is suggested that the proper selection of solvent for a measurement in a given region of the spectrum, a concentration of protein providing an adequate signal for accurate measurement, and the proper selection of cell path length to yield optimal optical density and clarity of the solution are some considerations relevant to CD spectroscopic measurements. The proper manipulation of the scanning speed of the instrument, the time constant for response, and the required signal-to-noise ratio are interrelated features which must be understood through use of the instrument. The CD spectrum of a hemoprotein, as of other proteins, in the region below 250 nm is expressed in terms of the contribution from the amide bonds of the molecule as residue ellipticity, and has the dimensions of deg-cm2/decimole of amide bonds. The observed quantity is transferred to residue ellipticity. The analysis of CD spectra of proteins in terms of the conformation of the chromophores is accomplished through two broad approaches: the theoretical approach and the phenomenological approach.