Section:General Biochemistry » Bioenergetics

ATP Synthesis Atp Synthesis in Ch

loroplasts is called photophosphorylation and is similar to oxidative phosphorylation in mitochondria. The light-driven transport of electrons from water to NADP+ is coupled to the translocation of protons from the stroma across the thylakoid membrane (the green, energy-converting membrane) into the lumen. Electron transport from Q to P700+ is exergonic. Part of the energy released by electron transport is conserved by the formation of an electrochemical proton gradient. The cytochrome b6f complex of chloroplasts functions not only in electron transport, but also in proton translocation. The active site of the oxygen-evolving enzyme is arranged so that the protons formed during water oxidation are released into the thylakoid lumen. These protons contribute to the electrochemical proton potential. The thylakoid membrane contains a protein that functions to transport Cl across the membrane. Proton accumulation in the thylakoid lumen is electrically balanced in large part by Cl uptake. As a result, thylakoids accumulate HCl and the membrane potential across the membrane is low. The pH inside the lumen during steady-state photosynthesis is about 5.0.

One of the earliest experiments that supported the hypothesis that ATP synthesis and electron transport were linked by the electrochemical proton potential was carried out with isolated thylakoid membranes. Thylakoid membranes were placed in a buffer at pH 4.0 and after a few seconds the pH was rapidly increased to 8.0, which resulted in the formation of a proton activity gradient. This artificially formed gradient was shown to drive the synthesis of ATP from ADP and Pi. The experiments were carried out in the dark so that the possibility that electron transport contributed to the ATP synthesis was excluded. Thus, a proton activity gradient was proven capable of driving ATP synthesis.

The thylakoid membrane enzyme that couplesATP synthesis to the flow of protons down their electrochemical gradient is called the chloroplast ATP synthase (see Fig. 10). This enzyme has remarkable similarities to ATP synthases in mitochondria and certain bacteria. For example, the β subunits of the chloroplast ATP synthase have 76% amino acid sequence identity with the β subunits of the ATP synthase of the bacterium E. coli.

The reaction catalyzed by ATP synthases is:

⇒ Equation [11] nHa+ + ADP + Pi + H+ → nHb+ + ATP + H2O,

where n is the number of protons translocated per ATP synthesized, probably three or four, and a and b refer to the opposite sides of the coupling membrane. Provided the electrochemical proton potential is high, the reaction is poised in the direction of ATP synthesis. In principle, when the proton potential is low, ATP synthases should hydrolyze ATP and cause the pumping of protons across the membrane in the direction opposite that which occurs during ATP synthesis. ATP-dependent proton transport by theATP synthase is of physiological significance in E. coli under anaerobic conditions in that it generates the electrochemical proton potential across the plasma membrane of the bacterium. This potential is used for the active uptake of some carbohydrates and amino acids.

In contrast, ATP hydrolysis by the chloroplastATP synthase in the dark has no physiological role and would be wasteful. In fact, the rate of ATP hydrolysis by the ATP synthase in thylakoids in the dark is less than 1% of the rate of ATP synthesis in the light. Remarkably, within 10–20 msec after the initiation of illumination, ATP synthesis reaches its steady-state rate. Thus, the activity of the chloroplast ATP synthase is switched on in the light and off in the dark. In addition to being the driving force for ATP synthesis, the electrochemical proton potential is involved in switching the enzyme on. Structural perturbations of the enzyme induced by the proton potential overcome inhibitory interactions with bound ADP as well as with a polypeptide subunit of the synthase. An additional regulatory mechanism that is unique to the chloroplast ATP synthase is reductive activation. Reduction of a disulfide bond in a subunit of the chloroplast ATP synthase to a dithiol enhances the rate of ATP synthesis, especially at physiological values of the proton potential. The electrons for this reduction are derived from the chloroplast electron transport chain.