Jagendorf

The mechani.sm of photopho.sphorylation i.s chemio.smotic. In 1966, Jagendorf and Uribe experimentally demon.strated for the first time drat establishment of an electrochemical gradient across the membrane of an energy-tran.sdncing organelle conld lead to ATP. synthe.sis. They equilibrated isolated chloroplasts for 60 seconds in a pH 4 bath, adjusted die pH to 8 in die presence of ADP and Pi, and allowed phosphorylation to proceed for 15 seconds. The entire experiment was carried ont in tire dark. 

Jagendorf, A. T, and Uribe, E., 1966. ATP formadon caused by acid-base transidon of spinach chloroplasts. Proceedings of the National Academy of Sciences, USA 55 170-177. The classic paper providing the first experimental verificadon of Mitchell s chemiosmodc hypodiesis. 

Electron-transferring molecules in the chain of carriers connecting PSII and PSI are oriented asymmetrically in the thylakoid membrane, so photoinduced electron flow results in the net movement of protons across the membrane, from the stromal side to the thylakoid lumen (Fig. 19-57). In 1966 Andre Jagendorf showed that a pH gradient across the thylakoid membrane (alkaline outside) could furnish the driving force to generate ATP. 

Jagendorf, A.T. (1967) Acid-base transitions and phosphorylation by chloroplasts. Fed. Proc. 26, 1361-1369. 

Other optional experiments may be completed if time allows. For example, the effectiveness of various redox dyes may be analyzed. In addition to those listed in the text, FMN, ferricyanide, and dichlorophenolindophe-nol may be tested (Neumann and Jagendorf, 1964). It has been shown that NH4C1 and amines stimulate proton uptake. If a potassium ion-specific electrode is available, the light-induced efflux of K+ from spinach chloroplasts may be studied (Dilley, 1972). 

A. Jagendorf and E. Uribe, Proc. Natl. Acad. Sci. USA, 55, 170-177 (1966). ATP Formation Caused by Acid-Base Transition of Spinach Chloroplasts. ... 

J. Neumann and A. Jagendorf, Arch. Biochem. Biophys. 107, 109-119 (1964). Light-Induced pH Changes Related to Phosphorylation by Chloroplasts. ... 

Observations in chloroplasts played a key role in the development of the chemiosmotic theory of oxidative phosphorylation, which we discussed in chapter 14. Andre Jagendorf and his colleagues discovered that if chloroplasts are illuminated in the absence of ADP, they developed the capacity to form ATP when ADP was added later, after the light was turned off. The amount of ATP synthesized was much greater than the number of electron-transport assemblies in the thylakoid membranes, so the energy to drive the phosphorylation could not have been stored in an energized... 

JAGENDORF, A. T., TAKABE, T., Inducers of glycinebetaine synthesis in barley., Plant Physiol., 2001,127, 1827-1835. 

One of the most striking pieces of evidence in support of the chemiosmotic hypothesis was obtained in the 1960s by Andre Jagendorf and Ernest Uribe. When chloroplast lamellae are incubated in a solution at pH 4—in which case pH1 presumably attains a value near 4 — and then rapidly transferred to a solution with a pH° of 8 containing ADP and phosphate, the... 

In 1966, Andre Jagendorf showed that chloroplasts synthesize ATP in the dark when an artificial pH gradient is imposed across the thylakoid membrane. To create this transient pH gradient, he soaked chloroplasts in a pH 4 buffer for several hours and then rapidly mixed them with a pH 8 buffer containing ADP and Pj. The pH of the stroma suddenly increased to 8, whereas the pH of the thylakoid space remained at 4. A burst of ATP synthesis then accompanied the disappearance of the pH gradient across the thylakoid membrane (Figure 19.24). This incisive experiment was one of the first to unequivocally support the hypothesis put forth by Peter Mitchell that ATP synthesis is driven by proton-motive force. 

Figure 19.24. Jagendorf s Demonstration. Chloroplasts synthesize ATP after the imposition of a pH gradient.

The respiratory and photosynthetic electron-transfer pathways are proton pumps operating with the same polarity as does the A TP synthase when hydrolyzing A TP Since it is difficult to detect protons circulating in the steady-state, Mitchell and Moyle [19] studied the transient extrusion of protons when a small amount of oxygen is injected into an anaerobic incubation of mitochondria in the presence of substrate. Prior to this, Neumann and Jagendorf [20] had observed a light-dependent proton uptake into chloroplast thylakoid membranes. 

Direct evidence for the chemiosmotic theory [1] is proton motive ATP synthesis in purified FgFj proteoliposomes on applying [38,131]. Jagendorf and Uribe [132] first demonstrated ATP synthesis in subchloroplast particles that had been loaded with a weak acid (pH 4.5) and were then transferred to alkaline media at pH values... 

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