Cytochrome b /// the Respiratory Chain 595 



of oxygen, caused by the aerobic oxidation of succinate. In our view, the course of 

 the reduction suggests that, after a very short time lag, cytochrome t> is as rapidly 

 reduced as cytochrome rt,. If this interpretation is correct, it follows that cyanide 

 must be interfering with the reduction of cytochrome b. One possible way in which it 

 might do this is suggested in the paper. 



Whether azide, U.,S or CO have the same effect as cyanide has not yet been tested. 

 Antimycin clearly does not. Chance's report, which I do not think has been mentioned 

 in his papers, that the reduction of cytochrome b by liver mitochondria is rapid in the 

 presence of cyanide, is another interesting example of the fact, first found by Chance, 

 that the kinetics of cytochrome b are different in phosphorylating and non-phosphory- 

 lating preparations. This is one of the points which we have attempted to explain by 

 the speculations at the end of our paper. 



Figure 3 (Slater and Colpa-Boonstra, this volume, p. 582) shows, in fact, that 

 reduction of cytochrome b begins a short time (about 0-25 sec) after that of cyto- 

 chrome 03. But this is to be expected if cytochrome b reacts at the substrate end of the 

 chain, and cytochrome a^ at the oxygen end. The important point, to our mind, is that 

 after this initial lag, the reduction of cytochrome b follows the same course as that of 

 cytochrome 03. This rapid reduction of cytochrome b has been found by Colpa- 

 Boonstra both with our recorder in Amsterdam, and with that of Holton which has a 

 faster response time. The possibility that the limiting factor in the speed of the tracing 

 pen was the response time of the instrument was excluded after a careful examination 

 of this point by Colpa-Boonstra. 



The haem which, as measured by the intensity of the pyridine haemochrome 

 band, disappears on treatment of a heart-muscle preparation with BAL in the presence 

 of air is probably largely accounted for by the destruction of the myoglobin and 

 cytochrome b' which are present in the preparation. This destruction cannot be 

 related to the complete inactivation of the respiratory chain which also results from 

 this BAL treatment, unless cytochrome b' is in some way involved in the chain. This 

 appears to us rather unlikely, although not impossible, in view of Chance's finding that 

 in the presence of antimycin, cytochrome b' is rapidly reduced by succinate and DPNH. 



Deul (1959) has evidence that, during treatment of haemoglobin with BAL, not 

 only is a methene bridge carbon atom attacked with formation of bile pigments, but 

 also one or both of the vinyl side chains are attacked, possibly with the formation 

 of a thio-ether similar to that found in cytochrome c. We are at present investigating 

 the working hypothesis that I2 in our proposed mechanism is one of the vinyl groups 

 of cytochrome b, and that this group is the site of action of both BAL and antimycin. 

 Chance: The position may be summed up in the following terms: Colpa-Boonstra and 

 Slater include cytochrome b in the non-phosphorylating succinate oxidase system at 

 room temperatures, but have reasons to doubt its participation in the chain in the 

 presence of cyanide. It is my opinion that the demonstrations at low temperatures 

 and in the presence of inhibitors, either cyanide or malonate, provide an affirmative 

 answer to the question as to whether electrons can be transferred to cytochrome c 

 without having passed through cytochrome b. The question of whether cytochrome 

 b can ever act fast enough to transfer electrons to cytochrome c appears to be answered 

 affirmatively for phosphorylating preparations, and Colpa-Boonstra and Slater feel 

 that they have evidence that their non-phosphorylating preparations show electron 

 transfer function of cytochrome b at room temperature. It is also true that various 

 types of electron transfer particles obtained from Green's laboratory show varied 

 responses of cytochrome b (Chance, unpublished data). 



It is of some importance to determine whether three conditions for cytochrome b 

 activity need be postulated: (1) full participation in phosphorylating systems, (2) 

 full participation in non-phosphorylating systems, (3) no participation in non-phos- 

 phorylating systems. It is possible that there are some vestiges of energy conservation 

 remaining in some heart muscle preparations that make it necessary to consider 

 mechanisms only for the first and third conditions. 

 Slater: The question which has interested us for some years is whether cytochrome 

 b is on the main chain when succinate or DPNH is oxidized by non-phosphorylating 



