58 1. MALONATE 



rates of penetration of succinate and malonate into cells, obtained prefer- 

 ably with radioactive material and under the same conditions. Quantitative 

 studies on the malonate inhibition of intracellular dye reduction resulting 

 from succinate oxidation might also be informative in certain instances. 

 It has been shown that malonate inhibits the succinate-induced reduction of 

 neotetrazolium in adipose tissue cells (Fried and Antopol, 1957), but noth- 

 ing otherwise is known about the succinate dehydrogenase from this tissue. 



Competitive Nature of the Inhibition in Cellular Systems 



It has been shown in several tissues that the addition of succinate will 

 reverse the inhibition of respiration produced by malonate. Thus in Avena 

 coleoptile the inhibition by 50 mM malonate is reduced from 57.4% to 

 25.8% upon adding succinate (Bonner, 1948), and in spinach leaves from 

 75.4% to 20.5% (Bonner and Wildman, 1946). In chick embryonic carti- 

 lage, the depression of respiration by 10 mM malonate is reversed by 100 

 mM succinate but not by 10 mM (Boyd and Neuman, 1954). Such results 

 have occasionally been stated to prove the competitive nature of the inhibi- 

 tion but this reasoning is not completely valid. The mere increase in O2 up- 

 take seen on addition of succinate to malonate-inhibited tissues is alone 

 not evidence for competition. The effects of succinate on uninhibited tissue 

 must also be tested and it must be shown that the actual inhibition is de- 

 creased. A decrease in the inhibition brought about by increasing succinate 

 concentrations has indeed been reported in two tissues, pigeon breast muscle 

 (Krebs and Johnson, 1948) and the trypanosome Crithidia (Hunter, 1960) 

 and in the latter a true competitive inhibition was demonstrated by 1/v — 1 /(S) 

 plots. The data are given in Table 1-10. It is probable that the inhibition 

 of succinate oxidation in cellular systems by malonate would frequently 

 not obey strictly competitive kinetics, due to the various complexities that 

 arise, as discussed in the previous section, even though the primary inhi- 

 bition on the succinate dehydrogenase were competitive. Some of the prob- 

 lems involved in the determination of the type of inhibition in cells have 

 been discussed in Chapter 1-9. 



INHIBITIONS OF ENZYMES 

 OTHER THAN SUCCINATE DEHYDROGENASE 



It is very important to establish the degree of specificity that may be 

 achieved in the use of malonate under various conditions. To this end 

 we shall first discuss the direct evidence for the inhibition of enzymes 

 other than succinate dehydrogenase, and then proceed to the effects on the 

 operation of the tricarboxylic acid cycle, the accumulation of succinate and 

 other intermediates, and finally the antagonism of the malonate inhibition 



