Heme proteins myoglobin reconstitution

One of the variables in the structures of the porphyrins present in heme proteins is the presence or absence of vinyl substituents on the periphery of the macrocycle. For example, b hemes have vinyl substituents whereas c hemes do not. Because of the sensitivity of such vinyl substituents during synthetic transformations, it has often been desirable to use octa-alkyl porphyrins in model studies of the spectroscopic properties of heme systems. The development of improved methods for the preparation of octa-alkyl porphyrins has likewise increased the availability of such porphyrins for model studies (20, 21). To assess the effect that replacement of the two vinyl substituents in protoporphyrin IX with alkyl (ethyl) groups has on the MCD properties of the heme system, an extensive and systematic study of the MCD properties of mesoheme IX-reconstituted myoglobin and horseradish peroxidase in comparison with the spectra of the native protoheme-bound proteins has been carried out (22). The structures of these two porphyrins are shown in Figure 3. 

CD can be used to monitor the phenomenon of heme isomerism in heme proteins. " The isomerism, discovered by LaMar and co-workers, results from the presence of two forms of myoglobin which differ by 180° in the heme orientation about the a, y-methine carbon axis. The two isomers are present in a 9 1 ratio at equilibrium in carbonmonoxymyo-globin (MbCO) but are in a 1 1 ratio in a sample freshly reconstituted from apomyoglobin and heme. Freshly reconstituted MbCO has a Soret CD band with only about half the amplitude of the native form, " suggesting that the minor isomer (at equilibrium) has only a weak Soret CD spectrum. The major form has a strong positive Soret CD band, whereas the minor form has a weakly negative Soret CD band. These results and the CD of the heme undecapeptide from cytochrome indicate that the origin of the Soret CD band must be more complex than proposed by Hsu and Woody, ... 

---