70 1. lODOACETATE AND lODOACETAMIDE 



with the inhibitions of the EM pathway (Table 1-11) because of the dif- 

 ferent conditions under which the experiments were run, but it is evident 

 that the sensitivities are roughly the same. It is not necessary that they 

 be the same, even though all the inhibition is on 3-PGDH, since in such a 

 complex multienzyme system involving phosphorylations the inhibition on 

 the whole system may be more than on a single component enzyme. One 

 can only say that the EM pathway inhibitions observed can usually be 

 attributed to an action mainly, if not entirely, on 3-PGDH. A more im- 

 portant question is whether other enzymes in the EM pathway are inhib- 

 ited significantly at iodoacetate concentrations blocking the pathway over 

 90% (i.e., up to around 0.2 mM). Table 1-12 summarizes inhibitions on the 

 enzymes of the EM pathway. There is very little evidence that any enzyme 

 involved in the direct pathway is inhibited to an important degree at these 

 concentrations. Even the hexokinases, probably the most sensitive enzymes 

 other than 3-PGDH, apparently are not inhibited appreciably by 0.2 mM 

 iodoacetate, but certainly are when the concentration is 1 mM or over. 

 When higher concentrations of iodoacetate are used, one would expect the 

 phosphorylation of glucose to be depressed directly (and perhaps by de- 

 pletion of ATP indirectly). ^-Glycerophosphate dehydrogenase is fairly sen- 

 sitive also but is not on the direct pathway; however, inhibition of this en- 

 zyme may be important in the effects of iodoacetate on glycerol fermenta- 

 tion where dihydroxyacetone-P is reduced by NADH via this enzyme. One 

 may conclude that iodoacetate in reasonable concentrations can exert a 

 selective action on 3-PGDH with respect to the EM pathway. Turning to 

 the additional enzymes involved in anaerobic glycolysis (lactate dehydro- 

 genase. Table 1-5) and alcohol fermentation (alcohol dehydrogenase, Table 

 1-4), there is again little evidence that these enzymes are inhibited signif- 

 icantly by low iodoacetate concentrations, except possibly in some plants 

 (e.g., Avena alcohol dehydrogenase is inhibited 73% by 0.1 mM iodoace- 

 tate) and in yeast. Although the data are not conclusive, it appears that 

 alcohol fermentation in yeast must be inhibited by a combination of ac- 

 tions on 3-PGDH and alcohol dehydrogenase. Unfortunately we have no 

 reliable data on muscle lactate dehydrogenase, so it is impossible to evaluate 

 the importance of this enzyme in the inhibition of glycolysis in muscle. 



(b) Accumulation of intermediates during iodoacetate inhibition. Inhibition 

 of a step in a monolinear sequence of irreversible reactions leads to the ac- 

 cumulation of the intermediate which is the substrate for the blocked en- 

 zyme; but when the reactions are mainly reversible the situation will be 

 more complex and the intermediates accumulating may be those some dis- 

 tance proximal to the block, the relative concentrations being determined 

 in part by thermodynamic equilibria between the substances. The EM path- 

 way is of the latter type. Accumulation of 3-phosphoglyceraldehyde would 

 thus not be expected to be marked for two reasons. At equilibrium the ratio 



