Analysis and Interpretation of Absorption Spectra of Haemin Chromoproteins 171 



Drabkin: Williams has suggested that the band at 500 m/i, which is No. 5 of my frequency 

 distributed series and ascribed by me to originate in the dynamic haemin structure, is 

 rather owing to iron itself. Having earlier discussed this question with him, I believe 

 his deduction originates from finding a maximum at about 510 m/t in such complexes 

 as ferrous orthophenanthroline (see my Fig. 2). This is a very broad maximum and 

 doubtless includes several component bands. Of course my proposal also involves 

 a co-ordination complex. I must respect his theoretical knowledge, but naturally I 

 prefer my own proposal. At any rate, I believe we agree on many other aspects of 

 our individual analytical approaches. Thus, we have brought out somewhat similar 

 correlations between the absorption spectra and corresponding magnetometric data 

 (see my Table 3 and text, p. 158). 



It may be pointed out that, in Table 3, in the case of ferrihaemoglobin and ferri- 

 haemoglobin hydroxide, one may now substitute the terms 'high-spin' and 'mixture 

 of high-spin and low-spin' in place of the older 'essentially' or 'partially' ionic (see 

 Orgel's, Williams', and George's papers, this symposium). I wonder whether it is 

 possible that the unexpected behaviour in the vicinity of the Soret band (my band 

 No. 6) in the transition spectra of MHb -> MHbOH (i.e. the absence of an isosbestic 

 point) may be related to a change from a high-spin complex to a mixture of high- 

 and low-spin ? 



Wainio: I wish to ask Drabkin, if the a- and )5-bands of the haem of any one of the haem- 

 proteins is replaced by the bands of the corresponding porphyrin, will the latter fall 

 into the frequency distribution series? 



Drabkin: In the ultra-violet region they do not fit well. Since I was making the most 

 simplifying assumption, namely that the spectra are an expression of iron in a co- 

 ordinated complex, I was content to start with, and confine myself to iron-porphyrin 

 complexes. I am not at all convinced that there is real validity in drawing parallels 

 between porphyrin bands and correspondingly located bands of haemin derivatives. 



The bands in the region of 830 and 280 m/x 



George: I should like to ask Drabkin whether the near infra-red bands of the ferri- 

 myoglobin fluoride complex at 740 and 830 m/t can be fitted into the frequency series 

 with successive values of 'rt'. If the wave-number separation is too small then one or 

 the other would have to be regarded as belonging to another category like the a- and 

 /S-bands. 



Drabkin: The band at 830 m/i is present in other haemin protein derivatives (as evident 

 in Table 2 of my paper). This band corresponds to v x 10""^ = '-^120, and is No. 3 

 of my proposed equally-spaced frequency series. The band at 740 m/< may correspond 

 with the at present anomalous band at 760 m/z of deoxygenated haemoglobin (see my 

 Fig. 1), and may in fact belong to a different series, or may be 'odd-man-out' as you 

 have expressed it. Incidentally, in some derivatives (see Fig. 1, p. 143) there are 

 maxima in the vicinity of wavelength 920 m/n which do not fit in the main series. This 

 is brought out in my paper. 



George: For myoglobin derivatives the millimolar extinction coefficients in the protein 

 absorption region, 260-290 m/<, are about 30 compared to the value of about 13 for 

 apomyoglobin. Similar values have been reported for haemoglobin derivatives and 

 apohaemoglobin, hence there can be no doubt that the haem absorbs significantly in 

 this region. The same is true of vitamin B12 and free benzimidazole, so the eff'ect is 

 not restricted to haemoproteins. 



Margoliash: In the case of cytochrome c, for which the amino acid composition is 

 reasonably well known, it is easy to calculate the contribution of tryptophane, phenyl- 

 alanine and tyrosine to the 280 m/i band. The results are roughly the same as those 

 quoted by George for metmyoglobin. There is, moreover, a striking difference between 

 the spectra of the ferro- and ferri- forms of cytochrome c in the 'protein' band region, 

 indicating a distinct contribution of haem absorption to this band (Margoliash and 

 Frohwirt, Biocliem. J. 71, 570, 1959). 



H.E. — ^VOL. I — N 



