592 



Lester Packer 



as PMS and Mg"^ ("^1/2 ^°^ activation = 70 sees). In view of this 

 it was of interest to examine the action of ATP on chloroplast 

 structure in the dark, following hrief periods of pre- illumination. 

 As sho-vm in figure ^1-, pre- illumination of chloroplasts in the 

 presence of the complete system with ATP, even after extremely 

 short light intervals, resulted in a pronounced effect of ATP on 

 chloroplast structure in the dark. Eventually, after 60 sees pre- 

 illumination, the light- scattering first decreases, and then in- 

 creases in the dark; thus arriving finally at the condition ob- 

 served after the more prolonged 5 min illvimination (cf. figure 3) - 



To define more precisely the action of ATP, an experimental 

 design was developed to permit the simultaneous measurement of its 

 hydrolysis measured by pH change with light- scattering. Control 

 experiments showed a linear relationship between the extent of pH 

 decline and ATP expenditui'e. Calcul.ations of the rate of ATP 

 hydrolysis made by the pH method closely corresponded with chemi- 

 cal analysis. In view of this, it was possible to follow the time 

 course of ATP hydrolysis with light- scattering changes. Figure 5 

 shows the typicaj. light- scattering responses under conditions of 

 ATP hydrolysis. The pH trace indicates the requirement for pre- 

 illuiTiination for activation of ATP hydrolysis. After removal of 

 actinic light, hydrolysis of ATP proceeds in a linear fashion and 

 an incomplete reversal of the scattering response characteristic 

 of these conditions is observed. Upon exhaustion of ATP in the 

 dark as indicated by the cessation of the pH change, a complete 

 reversal of the scattering level to the original dark level is 

 observed. This experimeiit provides proof that ATP and its hydro- 

 lysis are responsible for maintenance of the steady state level of 

 scattering change observed in the dark. 



It was therefore of interest to ascertain the action of ATP on 

 the steady state of scattering under conditions favorable for both 

 synthesis and hydrolysis. This was tested under conditions of 

 continuous Illumination and it was found that the scattering 

 change was U8fj above the dark control. Upon exhaustion of ATP in 

 the lig).it, a decline of the steady state of light scattering 

 change to '^Q'fo could be observed. 



In view of the close correlation of the action of ATP and its 

 hydrolysis to conditions necessary for generating scattering 

 changes in chloroplasts, it seemed possible that the action of ATP 

 might be associated with the presence of some contractile princi- 

 ple. Accordingly, chloroplasts were extracted under the same 

 conditions employed for the extraction of actomyosin from mamma- 

 lian muscle. A protein fraction could be extracted from chloro- 

 plast membranes which hydrolysed ATP and ITP. The fraction also 

 showed decreases in light- scattering on addition of these nucleo- 



