STUDIES ON THE MECHANISM OF PHOTOPHOSPHORYLATION 

 A. T. Jagendorf and Geoffrey Hind 



Our search for insight into the mechanism of ATP 

 formation by chloroplasts has been based, in the recent 

 past, on a two stage technique. The principle is to 

 illuminate chloroplasts together with some, but not all, 

 the components necessary for phosphorylation of ADP; 

 then turn off the light and add the missing compounds. 

 Such a procedure should be capable of providing 

 information on the existence and survival time of 

 intermediates between the initial light absorption, 

 and the final pyrophosphate bond. 



The first attempts with this approach were 

 interpreted as showing the existence of a high energy 

 phosphorylated intermediate (l). Results with improved 

 techniques (cutting the time interval between 

 illumination and dark additions to a fraction of a 

 second, by illuminating in a syringe, and injecting 

 directly into a completely dark test tube) now lead us 

 to believe that the earlier interpretation was probably 

 not correct. The earlier results may be explained 

 instead by something unsuspected at that time: the 

 existence of a light-induced transient ATPase which is 

 strongly inhibited by ADP at concentrations as low as 

 20 uM. This phenomenon is distinct from the two 

 reported ATPases (^,5,6) in certain of its requirements 

 and should be helpful in the elucidation of 

 photophosphorylation from the reverse direction. 



The more rapid injection technique has shown that 

 a non-phosphorylated intermediate (or intermediate state) 

 of the chloroplasts does exist, and in major amounts 

 under the proper circumstances. Although it has not 

 been identified, we have found a correlation of the 

 high energy state of the chloroplasts with both a very 

 major pH change, and with a non-specific alteration in 

 the optical density, probably representing a change in 

 chloroplast geometry. 



Light activated ATP breakdown. 



Table 1 shows the fate of the charcoal adsorbable 

 counts formed by illuminating chloroplasts with P , and 

 no added ADP. Two features - the loss of counts in the 



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