EFFECTS ON TRICAKBOXYLIC ACID CYCLE 69 



all. The examples discussed above indicate the impossibility of establishing 

 such a mechanism without considering the changes in malonate concentra- 

 tion or treating the data quantitatively. Despite this lack of positive evi- 

 dence, there is certainly no doubt but that this type of inhibition can occur 

 and may be sometimes very important. In the oxidation of pyruvate by 

 rat heart mitochondria, the Mg++ concentration must fall below 1 mM 

 before there is any significant decrease in the rate (Montgomery and Webb, 

 1956 b). We usually used 5 mM Mg++ in the medium so that it would have 

 required at least 40 mM malonate to produce a detectable inhibition by this 

 mechanism. Malonate at 50 mM did indeed inhibit around 50% but this 

 must certainly be due to other actions to a large extent. These experiments 

 were done with the a-ketoglutarate oxidase blocked by parapyruvate so 

 that any inhibition of succinate oxidation would not be involved. 



EFFECTS OF MALONATE ON THE OPERATION OF THE 

 TRICARBOXYLIC ACID CYCLE 



Malonate is usually assumed to produce its major effects on cellular meta- 

 bolism and function by disturbing the operation of the cycle* and reducing 

 the rate of formation of ATP. Malonate has often been used to establish if 

 the cycle is operative in a tissue or if a particular functional activity is 

 dependent on the cycle. It is thus important to examine critically the 

 nature of the cycle block and the effects it may have on the over-all oxida- 

 tive metabolism. There are two aspects that are especially relevant to this 

 question. There is the problem of the specificity of action of malonate on 

 succinate dehydrogenase and this will be considered later. In the present 

 section we shall assume that the only inhibition is on the oxidation of suc- 

 cinate and discuss the problems relative to the interpretation of such 

 a block. Before treating the actual results obtained with malonate, the 

 nature of the cycle and its responses to inhibition will be outlined. 



Some General Principles of Cycle Block 



The primary function of the cycle is to incorporate and oxidize acetyl-CoA, 

 whether this arises from pyruvate, acetate, fatty acids, or elsewhere, and 

 thus it is particularly important to discuss the effects of malonate block on 

 this. The situation is relatively clear in suspensions of isolated mitochondria, 

 in which the concentrations of the cycle intermediates are low and the endo- 

 genous respiration is generally negligible. Pyruvate, or other substances 

 giving rise to acetyl-CoA, may be oxidized through the cycle only if some 



* In this chapter the term "cycle" will always refer to the tricarboxylic acid cycle 

 for convenience and other cycles will be designated by their special names. 



