Acid-to-Base Phosphorylation as a Measure of Plastid Membrane Maturity

Acid-to-Base Phosphorylation as a Measure of Plastid Membrane Maturity 

What other light-independent partial reactions of photosynthesis could we seek in maize etioplasts Jagendorf and Uribe demonstrated that thylakoids of spinach form ATP from ADP and inorganic phosphate when they are transferred from an acidic to an alkaline solution. The formation of ATP during acid-to-base transition (A/B) requires no light. In fact, the reaction is generally carried out in the presence of p-chlorophenyl-1,1-dimethyl urea (CMU), a powerful inhibitor of photosynthesis, to prevent photophosphoiylation. 

The assay is relatively simple. A thylakoid preparation is incubated at about pH 4 in a CMU-containing succinic acid solution at O C for 30 sec. It is then mixed with a buffered solution (pH 8.15) containing ATP and Na2H P04. After 15 sec, trichloroacetic acid is added to terminate the reaction. The ATP formed is isolated and counted. The amount of Pi esterified is a measure of the rate or extent of the reaction. 

For one, the coupling factor for photophosphorylation is needed for A/B phosphorylation. This was shown by stripping green membranes with 

01 M NaCl-0.15 mM EDTA and collecting the membranes by centrifugation. The stripped membranes could not carry on A/B phosphorylation. The capacity to do this was, however, restored by incubating coupling-factor preparations from chloropiasts or etioplasts of maize with stripped thylakoids in 0.01 m EDTA-0.1 m MgCl2. This showed that the CFi was required, but was probably not the limiting factor since it is present in etioplasts. What other conditions are required  

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