INHIBITION OF SUCCINATE DEHYDKOGENASE 41 



kamp and Hill, 1951), malondiamide (Fawaz and Fawaz, 1954), and var- 

 ious derivatives of succinate, in which one or both of the carboxylate 

 groups have been replaced with nonionic groups (Dietrich et al., 1952), 



Malondialdehyde Malondiamide /3-Bromopropionate /3- Hydroxy propiononitrile 



are all lacking in inhibitory activity. An SH group close to the cationic 

 attachment points is proved by the high sensitivity of the dehydrogenase to 

 substances reacting with SH groups and the protection that malonate af- 

 fords against such substances. The latter is actually better evidence for the 

 proximity of the SH group because sulfhydryl reagents can alter protein 

 structure and exert effects for some distance over the enzyme, whereas the 

 blockade of these substances by a small molecule such as malonate would 

 be almost certain proof. Hopkins et al. (1938) showed that malonate, can 

 protect succinate dehydrogenase from oxidized glutathione (GSSG), which 

 is a potent inhibitor. Incubation of the enzyme with GSSG increased the 

 methylene blue reduction time from 10 min to 3 hr; malonate at concentra- 

 tions from 0.2 to 100 mM gave almost complete protection. That the 

 SH groups are protected by malonate was also shown by titration of these 

 groups with iodine. Potter and DuBois (1943) reported protection against 

 quinone inhibition and Barron and Singer (1945) against arsenicals. It is 

 interesting that malonate also protects against oxygen poisoning of suc- 

 cinate oxidase, which was interpreted to mean that the SH groups of the 

 enzyme are involved in this inactivation (Dickens, 1946 b). Oxalacetate 

 has also been shown to be protective against mercurials and arsenicals 

 (Stoppani and Brignone, 1957), providing evidence that it binds to the 

 same site as succinate and malonate. The degree of protection depends on 

 several factors, including the concentrations of the inhibitors and the rates 

 at which they react with the enzyme; the less protection seen against the 

 mercurials is attributed to their comparatively rapid action whereas the 



