Discussion 298 



King: In the system with ammonium salt and glucose, have you 

 observed any net growth of yeast? 



Holzer: Not in our experiments, because we only looked at what 

 happened after a few minutes; but after 30 to 60 minutes one can 

 observe growth. 



Chance : We never tried to observe gTowth. 



King: Then I do not understand how you define the steady state there. 



Backer: Perhaps you could tell us how you define growth? 



King: Growi:h is the net increase in cell mass. 



Holzer : If you wait for 30 minutes, you may get an increase of 3 to 5 

 per cent, and if you wait longer you may get the well known exponential 

 growth. What we have studied is what happens in the first few seconds 

 and minutes, and from this we hope to deduce how growth is initiated 

 by metabolic alterations. Therefore, our interest is focused on what 

 happens before you see an increase in cell mass. 



King : If we use a bacteriological term, then your study is still at the 

 lag phase? 



Holzer: Yes, that is right. 



Hinshelwood: When you add the ammonium salt to a yeast prepara- 

 tion, one of the most important points biologically is that the system is 

 preparing for synthesis, gro\\i:h and division. After some finite delay, 

 mass will increase, and division will set in. The two will not be phased 

 exactly together. It would be extremely interesting if these experi- 

 ments were carried to the stage where the first mass increase is observed 

 so that one could see which of the processes reaches a critical level and 

 triggers off something else. Presumably all the processes have been 

 evolved so as to fit cells optimally to grow and form new material. It is 

 interesting to note that a bacterial suspension which is reducing nitrate 

 vigorously, is completely inhibited for a long period if ammonium 

 salts are added. The explanation is fairly simple, and is an interesting 

 example of regulation: in the presence of ammonia, a much more 

 effective process goes on, which consumes the carriers much more 

 rapidly and reduces their concentrations to a level where they cannot 

 reduce nitrate. 



Magasanik : To introduce a note of caution, as far as the experiments 

 with ammonium salts are concerned: we have just by chance come 

 across two DPN-linked dehydrogenases, which require ammonium ion 

 for activity. One is inosinic acid dehydrogenase, where the ammonium 

 ion can be replaced by potassium. The other one is a glycerol dehydro- 

 genase in Aerobacter aerogenes, where ammonium ion is much more 

 effective than potassium. The affinity is low ; in other words, ammonium 

 salt has to be added to a high level. The ammonium ion may thus act 

 as an activator for dehydrogenase which in turn may bring about 

 changes in DPN levels. 



Lynen : To go back to the pH effect. Prof. Chance, did you add free 

 ammonia or ammonium chloride? 



Chance : Ammonium chloride. Dr. Magasanik, is the effect which you 

 describe reversed by hydrogen ions? 



Magasanik : No. 



