444 J- S. C. WESSELS 



TABLE I 



DiNITROPHENOL AS A CATALYST OF CYCLIC PhOTOPHOSPHORYLATION 



. jj. . , , s umoles ATP generated during 



Additions (umoles) ^ . ^,, . . * 



30-min. illumination 



0-5 vitamin K3 14 '4 



0-5 FMN 7-7 



3 DNP 2-5 



1-5 DNP 7-6 



0-6DNP 107 



03 DNP 79 



015 DNP 2-9 



No addition o-6 



o • 6 DNP ; dark o • 5 



0-6DNP 8-6 



0-6DNP + 30KCN 7-7 



o-6DNP + 3NaN3 69 



06 DNP + 3 NH.,OH 09 



06 DNP + 0012 CMU 2-9 



06 DNP + o-3 dicoumarol 3-5 



0-6 DNP + o-3 o-phenanthroline 1-7 



0-6 DNP + o • 3 /)-chloromercuribenzoate 2-5 



In addition, the reaction mixture included 40 /xmoles Na and K phosphate 

 buffer, pH 7 5; 10 /xmoles MgCL; 125 /imoles glucose; i ^umole ADP; 25 K.M. 

 units of hexokinase ; i ml. of a suspension of chloroplasts in o- 1 M tris (hydroxy- 

 methyl) aminomethane buffer pH 7-5, containing 0-5 mg. chlorophyll; and de- 

 ionized water to give a final volume of 3 -o ml. The reaction was carried out under 

 anaerobic conditions in Warburg manometer vessels as described previously [i]. 



A number of phenols have been tested for their catalytic activity in the 

 process of cyclic photophosphorylation and the results are given in Table II. 

 It seems that the presence of a nitro group is necessary though not sufficient 

 for the activity of the phenol derivatives. 



In order to act as an intermediate electron carrier across some gap in 

 the electron transport chain of isolated chloroplasts, DNP should be 

 transformed by chloroplasts into some reversible oxidation-reduction 

 system. Actually it was found that illuminated chloroplasts are capable of 

 reducing DNP quantitatively to 2-amino-4-nitrophenol, and that the latter 

 compound can serve as a cofactor for cyclic photophosphorylation. 



The reduction of DNP is dependent on light and on anaerobic condi- 

 tions. When the mixture is kept in the dark or when the chloroplasts are 

 illuminated in air, DNP can be recovered nearly quantitatively even after 

 several hours. This is also the case when boiled chloroplasts are illuminated 

 in the presence of DNP. The transformation of DNP into amino-nitro- 

 phenol proceeds approximately twice as fast if phosphorylating reagents 



