Figure 6.6 An Experiment Demonstrates the Chemiosmotic Mechanism
Previous research into the force driving ATP synthesis in mitochondria and chloroplasts led to the postulation of the chemiosmotic mechanism. Precisely how this mechanism uses ATP synthase to couple proton diffusion to ATP synthesis was demonstrated by two insightful experiments. In the first experiment (shown here), isolated organelles were placed in a medium with a low H+ concentration until equilibrium was reached. The organelles were then moved to a medium with a high H+ concentration. The H+ ions moved down the concentration gradient and into the organelles, driving the synthesis of ATP. In a second experiment, a proton pump was added to an artificial lipid vesicle and used to pump H+ into the vesicle, generating a gradient. ATP synthase was then inserted into the vesicle membrane. As the H+ diffused out of the vesicle, ATP synthase drove the synthesis of ATP. Together, the results of these two experiments provided evidence that the chemiosmotic mechanism uses ATP synthase to couple proton diffusion to ATP synthesis (see also Figure 6.5). It is important to realize that while ATP synthase normally functions to drive the synthesis of ATP, it is also capable of functioning in the reverse direction, leading to ATP hydrolysis and the formation of ADP and Pi. It is the H+ gradient, as well as the low concentration of ATP in the mitochondrial matrix, that drives the reaction toward ATP production. Consider, then, what would happen in the second experiment if a second ATP synthase, oriented in the opposite direction of the one originally inserted in the membrane, was added. In this scenario, the conditions for the second ATP synthase would differ from that for the original ATP synthase. The second ATP synthase would lack a H+ concentration gradient, thereby preventing ATP production. Further, the ATP produced by the first ATP synthase would likely drive the ATP synthesis reaction in the reverse direction, causing the enzyme to function as an ATPase and result in ATP hydrolysis.
Original Papers
Dr. Peter Mitchell received the Nobel Prize in chemistry in 1978 for the chemiosmotic theory of biological energy transfer.
Mitchell, P. 1961. Coupling of Phosphorylation to Electron and Hydrogen Transfer by a Chemi-Osmotic type of Mechanism. Nature 191: 144–148.
http://dx.doi.org/10.1038/191144a0
The original experiments on pH and ATP formation were done on chloroplasts, where the chemiosmotic generation of ATP is similar to mitochondria.
Jagendorf, A. T., and E. Uribe. 1966. ATP formation caused by acid-base transition of spinach chloroplasts. Proceedings of the National Academy of Sciences 55: 170–177.
http://www.pnas.org/cgi/reprint/55/1/170
Various experiments were repeated in mitochondria, with Dr. Paul Boyer later receiving the 1997 Nobel Prize in chemistry for research elucidating the importance of ATP synthase. See summary of the research experiments provided by:
Weber, M. 2002. Theory testing in experimental biology: the chemiosmotic mechanism of ATP synthesis. Studies in History and Philosophy of Science Part C: Studies in History and Philosophy of Biological and Biomedical Sciences 33: 29–52.
http://dx.doi.org/10.1016/S1369-8486(01)00016-4
Boyer, P. D., B. Chance, L. Ernster, P. Mitchell, E. Racker, and E. C. Slater. 1977. Oxidative Phosphorylation and Photophosphorylation. Annual Review of Biochemistry 46: 955–1026.
http://arjournals.annualreviews.org/doi/abs/10.1146/annurev.bi.46.070177.004515
Boyer, P. D. 2002. A Research Journey with ATP Synthase. Journal of Biological Chemistry 277: 39045–39061.
http://dx.doi.org/10.1074/jbc.X200001200
Links
Kimball’s Biology Pages: Cellular Respiration
http://users.rcn.com/jkimball.ma.ultranet/BiologyPages/C/CellularRespiration.html
Lane, N. 2010. Why Are Cells Powered by Proton Gradients? Nature Education 3(9): 18.
http://www.nature.com/scitable/topicpage/why-are-cells-powered-by-proton-gradients-14373960
Molecular and Cellular Biology Learning Center: Virtual Cell Animation Collection: Cellular Processes: ATP Synthase
http://vcell.ndsu.nodak.edu/animations/atpgradient/index.htm
University of California: Animation Movies of ATP Synthase
http://users.soe.ucsc.edu/~hongwang/Project/ATP_synthase/
Dr. Peter Mitchell: Nobel Prize in chemistry 1978
http://nobelprize.org/nobel_prizes/chemistry/laureates/1978/press.html
Dr. Paul Boyer: Nobel Prize in chemistry 1997
http://nobelprize.org/nobel_prizes/chemistry/laureates/1997/