ATP FORMATION BY SPINACH CHLOROPLASTS 459 



Mr. L. Choules and Dr. R. B. Ballentine at Johns Hopkins [lo]) the activity 

 appears to be associated with the major component. 



The enzvme characteristics known so far are compatible with a role 

 in phosphorylation. The pH optimum is a broad one, around 7 to 8. Mg 

 ions are needed, with 50",, activity at about 5 x io~^ m. Mg is replaceable 

 by Mn, Fe or Co ions. Although Ca has some low activity by itself, it is 

 inhibitory when combined with Mg. Zn, Hg, and Cu are inhibitors. 

 Ammonium ions are weak inhibitors also, with 50",, inhibition occurring 

 at I X io~^ M. Dinitrophenol has \irtually no effect. 



Efforts to show an irrele\ant acti\ity for this enzyme have so far failed. 

 There is no ATP--'^-P| exchange, no myokinase activity, no pyrophosphatase 

 or RNAase, no phosphatase for glucose-6-phosphate, fructose-6-phos- 

 phate, fructose- 1, 6-diphosphate, phosphoglyceric acid or phospho- 

 glvcerol. We can rule out glutamine synthetase, protein phosphokinase 

 (either with phosvitin or casein as substrate) and polynucleotide phos- 

 phorvlase. Xo transfer of high-energy phosphate to ADP occurs from 

 carbamvl phosphate, phosphoenolpyruvate, acetylphosphate, phospho- 

 serine or phosphocreatine. Other dinucleotides serve only weakly as 

 phosphate acceptors from ATP: GDP at 9",, of the rate of ADP, CDP 5"o, 

 UDP 19",,, and I DP 14",,. No products are formed other than ADP or 

 ATP in the usual reaction, and no other compounds (aside from Alg ions) 

 need to be added even to the highly dialyzed enzyme. 



In short, there is no sign of any reaction except for the exchange of a 

 high-energy phosphate from ATP to ADP. There is a very real possibility 

 that the reaction proceeds through a high-energy phosphate on the enzyme : 



ATP + E ^ E - P + ADP (4) 



If this is the case, the E ^ P might be related to or a part of the X ^ P 

 shown in the pre-loading experiments. Our future efforts will be devoted 

 to testing these aspects of the problem. 



References 



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2. .Avron, M., Krogmann, D. \\., and Jagendorf, A. T., Bioc/iitn. hiuphys. Acta 

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4. Jagendorf, A. T., Brookharoi Syvip. Biol. II, 236-258 (1958). 



5. Jagendorf, A. T., Fed. Proc. 18, 974-984 (1059). 



6. Avron, M., and Sharon, X., Biochevt. biophys. Res. Cnmm. 2, 336-339 (i960). 



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