Modification of the Secondary Structure of Haenioprotein Molecules 85 



Figure 4 shows the effect of addition of sodium sahcylate to haemoglobin. 

 The oxidase activity of haemoglobin increased with addition of low concentra- 

 tions of salicylate and reached its maximum at a salicylate concentration 

 which was not sufficient to cause an appreciable change of the absorption 

 spectrum of haemoglobin. A similar relation has been found with sodium 

 laurate instead of salicylate. Four moles of laurate/mole of haemoglobin 



100 



50 



01 0-2 0-3 0-4 0-5 



Salicylate, M 



0-6 



Fig. 4. Effect of salicylate on the oxidase activity of haemoglobin. The oxidase 

 activity was measured with a conventional Warburg technique in the presence of 

 ascorbic acid. The concentrations of haemoglobin and ascorbic acid were 9-4 X 

 10"^ M and 1 X 10"^ m, respectively. Ferrihaemochrome formation was measured 

 spectrophotometrically at 535 m/t. One hundred per cent promotion of oxidase 

 activity means that the oxidase activity with salicylate became twice as much as 

 that without salicylate. Curve I: promotion of oxidase activity; Curve II: 

 ferrihaemochrome formation. 



were sufficient to raise the oxidative activity to its maximum. It is evident 

 from this observation that the alteration of the secondary structure of the 

 protein part, required for the increased activity as an oxidative catalyst, has 

 already been completed at this concentration of laurate, although the modifica- 

 tion of the structure was not detectable spectroscopically and hence the 

 modification at this stage should be only slight. 



When the modification proceeds so far as to be spectroscopically detectable, 

 the appropriate condition for the activation of the latent oxidase activity of 

 the haemoglobin molecule is lost. It seems to be essential for activation of 

 the oxidase activity of haemoglobin that the extent of protein modification 

 should be slight. 



These experimental results are consistent with a stepwise nature of the 

 modification of the protein moiety of haemoglobin. Similar stepwise modifi- 

 cation was observed also with myoglobin and cytochrome c molecules 



