240 1. MALONATE 



Inasmuch as the active center of succinate dehydrogenase possesses a 

 sulfhydryl group close to the cationic binding sites, this being the basis 

 for the inhibition by mercurials and other sulfhydryl agents, the possibility 

 of combining a sulfhydryl inhibition with malonate presents itself. Mercuri- 

 malonamide and mercurimalonic diethyl ester have been prepared (Naik 

 and Patel, 1932) and these compounds, or particularly the hydrolyzed 



^COO-Et 

 ^C(X>-Et 



forms if they are stable, might be interesting to examine as inhibitors of 

 succinate dehydrogenase. 



Acetylene-dicarboxylate and Propane-tricarboxylate 



Succinate dehydrogenase is inhibited competitively by acetylene-dicar- 

 boxylate (Dietrich et at., 1952). The order of addition of the succinate and 

 acetylene-dicarboxylate is important; for example, if acetylene-dicarboxy- 

 late is added 20 min after the succinate, the rate of oxygen uptake de- 

 creases slowly and does not become equal to that observed when the suc- 

 cinate is added after the inhibitor until 5 hr (Thomson, 1959). Acetylene- 

 dicarboxylate is about as effective as malonate on long incubation with the 

 enzyme. The constants obtained on rat kidney succinate dehydrogenase 

 are ir,„ = 4.12 mM and K^ = 0.81 roM when substrate and inhibitor are 

 added together; after 18 hr incubation with the inhibitor, iC, =^ 0.171 vaM. 

 It is difficult to understand why the rate of inhibition is so slow. Succinate 

 dehydrogenase from pig heart is less readily inhibited, K^ being 1.4 mM 

 with A'^-methylphenazine as electron acceptor and 16.5 when ferricyanide is 

 the acceptor (Hellerman et al., 1960). Possibly insufficient time for equilib- 

 rium was allowed. The inhibitions of succinate and pyruvate oxidations 

 by acetylene-dicarboxylate in suspensions of rate heart mitochondria are 

 shown in Fig. 1-23 (Montgomery and Webb, 1956 b). Acetylene-dicarboxy- 

 late is less potent than malonate against succinate oxidation and more po- 

 tent against pyruvate oxidation, indicating that an inhibition is exerted 

 at some other point in the cycle. 



Propane-tricarboxylate is a rather weak inhibitor of succinate dehydro- 

 genase from rat heart, 5 mM inhibiting around 30% when the succinate is 

 also 5 mM. The oxidation of a-ketoglutarate is inhibited 60% under the 

 same conditions, suggesting some effect on the a-ketoglutarate oxidase. 

 The oxidation of pyruvate is inhibited even more strongly (Fig. 1-24). It 

 might be thought that propane-tricarboxylate would inhibit aconitase or 

 isocitrate dehydrogenase, but very little inhibition is noted, either of cit- 

 rate oxidation or the ability of citrate to function as a source of oxalac- 

 etate for the oxidation of pyruvate. 



