Studies on Mkrosonml Cytochroincs All 



TPNH, and the reduced isolated cytochrome is known to be capable of being oxidized 

 efficiently by cytochrome c, during the extraction procedure the reactivity towards 

 DPNH and TPNH is lost. It would be of great interest to know precisely on what the 

 TPNH reductase activity depends, for instance, whether a specific pyridine nucleotide 

 reductase such as that found for DPNH is involved, and the nature of its terminal 

 acceptor. Is it possible that, in the intact cell, the fact that the mitochondria lie in the 

 folds of the endoplasmic reticulum, which may therefore be in very close proximity to 

 mitochondria, could facilitate microsomal-mitochondrial interchange? However, 

 Chance's preceding remarks seem to make such an interaction improbable, since there 

 is little change in the oxidation-reduction level of cytochrome b^ in relation to changes 

 in that of other cytochromes. 



On the other hand the microsomes are the seat of an important series of hydroxyla- 

 tion, aromatization and ring-closure reactions which specificially require TPNH, and 

 might therefore represent a main route of TPNH oxidation in microsomes. My 

 colleagues, G. Clock and P. McLean, found for washed liver microsomes a ratio of 

 DPN+/DPNH of 5/1, suggesting that DPNH is probably capable of oxidation in micro- 

 somes, and since DPNH does not appear to be utilized there in the same pathways 

 as those which utilize TPNH, cytochrome b^ plus its specific cytochrome reductase 

 system would be suitable for DPNH oxidation provided some effective terminal 

 acceptor can be found. There remains, of course, the possibility raised by Chance's 

 comment, that alteration of the haemoprotein occurs during the extraction procedure 

 used. 

 Strittmatter : The possibility of microsomal-mitochondrial interchange to provide a 

 mechanism and possible function for the large pyridine nucleotide oxidizing capacity 

 of the microsomes is indeed attractive, but, as noted in my paper (p. 470), it finds no 

 direct support in the meagre data available. The observation of Chance that micro- 

 somal cytochrome in perfused liver slices or cell suspensions is not reduced under mild 

 reducing conditions, in contrast to the mitochondrial cytochrome components, also 

 militates against but does not rule out microsomal-mitochondrial interchange or a 

 respiratory role for cytochrome b^; reduction of cytochrome 65 requires a relatively 

 strong reducing agent, such as DPNH, which may be present in the intact cell of 

 solid liver but, as noted by Chance, may have been removed by his procedures for 

 preparing slices or cell suspensions. 



The Significance of E'q Values of Cytochromes in Relation to Cellular Function 



Estabrook: Strittmatter and Velick (/. biol. Cheni. 221, 265, 1956) have reported on the 

 oxidation-reduction potential of cytochrome 65. I wonder about the change in poten- 

 tial as determined with the particle bound haemaprotein (—0-120 V) and the purified 

 haemoprotein (+0-02 V), and the significance of this change in any consideration of 

 oxidation-reduction potential and the relevance to a thermodynamic consideration of 

 such potentials in energy transfer. 



Morton: There is a similar type of discrepancy, but in the opposite direction, in the 

 reported E'q values for soluble cytochrome b of heart-muscle (—005 V; Sekuzu and 

 Okunuki, /. Biochem. Tokyo 43, 109, 1956) and for cytochrome b in particulate 

 preparations (+0077 V; Colpa-Boonstra and Holton, Biochem. J. 11, iv, 1959). It 

 would appear that the reactivity of a cytochrome is altered by association with lipo- 

 protein material of cytoplasmic particles. 



Wainio: To demonstrate that the redox potential of an enzyme need not be different when 

 soluble and insoluble, we need only to consider the values obtamed for cytochrome c 

 oxidase by Ball (+0-29 V) who employed the heart muscle mitochondrial fragment, 

 and by Wainio (+0-285 V), who employed a deoxycholate-solubilized partially- 

 purified preparation of the oxidase. 



Strittmatter: As was noted in my paper (p. 464), the significance of this apparent change 

 in, potential on isolation of the particle-bound cytochrome cannot be assessed with 

 assurance at present. The change, if real, could reflect either the inevitable changes 

 in specific linkages, structural relations and physico-chemical milieu on separation 



