Analysis and Interpretation of Absorption Spectra of Haemin Chromoproteins 157 



spectral region would be four times higher than that of albumin (see curves 

 1 to 6 and the legend, Fig. 1 1). However, denaturation does have an influence 

 on chromoprotein spectra in this region, as it does on the albumin spectrum 

 (compare curves 4 and 7). The influence of haemin in this spectral region is 

 suggested by curves 4, 7 and 8. Contrariwise, if the protein moieties do exert 

 spectroscopic influence, they do so more evidently in the visible spectral 

 region, the spectral changes in which may be deduced to be a function of the 

 nature of co-ordinating groups or ligands bonded to the iron, which is also 

 bonded to the four pyrrolic nitrogens. Thus, the alkaline denaturation of 

 haemoglobin yields globin haemochrome, and the spectrum of globin ferro- 

 protoporphyrin in the visible spectral region is practically indistinguishable 

 from that of pyridine ferroprotoporphyrin (Drabkin, 1942a and Fig. 17). 

 The situation with respect to spectroscopically operative protein structures is 

 presumably different in the case of cytochrome b^. The pronounced maximum 

 at 265 m/i {v x 10^^ = 377) appears to be clearly ascribable to the riboflavin 

 phosphate and a polydeoxyribonucleotide, which are structural components 

 of this complex molecule (Appleby and Morton, 1954; Morton, 1958). 

 Whether the influence of the nucleotide is confined to this spectral region or 

 may be reflected in other spectral regions remains to be ascertained. Thus 

 far no convincing evidence has appeared that the number 9 band in the 

 chromoproteins may be a composite with finer structure attributable to the 

 aromatic amino acids, as has been shown for non-haemin proteins (Holiday, 

 1936,1937; Lavin, Northrop and Taylor, 1933; Lavin and Northrop, 1935). 

 It may be concluded that from the qualitative viewpoint the protein moiety of 

 the haemin proteins contributes only negligibly to their over-all spectra (see 

 Discussion). 



The a and (j Bands and the Neighbouring Visible Spectral Regions. Despite 

 the fundamental similarities disclosed by the analysis of the spectra of the 

 haemin proteins, in the past major attention has been devoted to characteristic 

 and pronounced diff'erences in these spectra in the visible spectral region. 

 Such diff'erences have been valuable in the accurate determination by means 

 of spectrophotometry of biologically important derivatives as well as in the 

 study of certain equilibria (Austin and Drabkin, 1935; Drabkin and Singer, 

 1939; Drabkin and Schmidt, 1945; Drabkin, 1950; Gordy and Drabkin, 

 1957), but may have directed attention away from the basic similarities. It 

 may be said that the combination of haemoglobin with gases (Og, CO, NO), 

 the oxidation of haemoglobin to ferrihaemoglobin, the alkaline denaturation 

 of haemoglobin to haemochrome, and the pH dependency of ferrihaemoglobin 

 are all spectroscopically operative, and that many of these reactions have 

 parallels in the spectroscopic behaviour of the cytochromes. Figures 12 to 17 

 and Table 3 are relevant in the interpretation of the diff'erences in the visible 

 spectra of chromoproteins, consonant with the analytical viewpoint that all the 

 spectra have a and /3 components straddled by bands 4 and 5 of the dominant 



