Form and Function Finally Prove Mitchell's Proton Motive Force

Volume 16 | Issue 13 | 28 | Jun. 24, 2002 Previous | Next Form and Function Finally Prove Mitchell's Proton Motive Force Perseverance, perspiration and a creative bent pay off | By Susan Jenkins The Faculty of 1000 is aWeb-based literature awareness tool published by BioMed Central. For more information visit www.facultyof1000.com. In 1961, Nobel laureate Peter Mitchell's provocative pairing of chemistry and biology gave birth to his chemiosmotic hypothesis and its corollary proton motive force, or what Mitchell liked to call proticity.1 Forty years later, the authors of these two selected Faculty of 1000 papers2,3 have provided structural and functional confirmation of Mitchell's theories. In the first paper, So Iwata and his graduate student Mika Jormakka provide molecular proof of proton motive force and an energy conserving redox loop,2 another Mitchell proposal. In the second manuscript, Andreas Matouschek and colleague Shihai Huang3 describe mitochondrial chemiosmosis and proton motive force in action. "It's a very, very old story," says Iwata, "and most of it came from Mitchell's imagination." Mitchell's theory, known as the chemiosmotic hypothesis, envisioned that positively charged hydrogen ions (protons) were transferred from one membrane side to the other, and that this process involved electron transfer reactions and adenosine triphosphate (ATP) synthesis. The theory explains how cells can generate different proton concentrations on a membrane's opposite sides and how this membrane potential can be used to provide energy for cellular processes.4 In his acceptance speech before the Nobel committee in 1978, Mitchell said, "Perhaps the most fruitful (and surprising) outcome of the development of the notion of chemiosmotic reaction is the experimental stimulus and guidance it has provided in work designed to answer" questions about cellular bioenergetics.5 Ten years after Mitchell won the chemistry prize, Nobelists Johann Deisenhofer, Robert Huber and Hartmut Michel received theirs for solving the first membrane protein structure. Iwata, one of Michel's postdocs, says his paper "shows that Peter Mitchell's original hypothesis of how cells convert energy into a usable form is correct." Iwata directs the Centre for Structural Biology at the Imperial College, UK. Iwata's group proves the molecular basis of proton motive force by crystallizing the structure of the membrane protein formate dehydrogenase-N from Escherichia coli. The 1.6Å-resolution structure demonstrates that a 90Å-long chain of 11 redox sites provides not only a continuous circuit through the enzyme, but also a direct connection to a menaquinone binding site. Furthermore, the structure clearly resolves the quinone-to-quinol cycle through nitrate reductase sitting on the opposite side of the membrane, completing a redox loop, also proposed by Mitchell. In his review, F1000 member David Richardson commented, "The enzyme structure, topology, and location of redox centers provide the first structural description of an electrogenic redox-loop mechanism for the generation of membrane potential." According to Iwata, membrane proteins, many of which are involved in genetic diseases, comprise about 30% of proteins in the human genome; of these, only three are eukaryotic. Iwata adds, "We [crystallographers] are solving about five to 10 membrane protein structures per year, which is very frustrating." In particular, his group is tackling G-protein-coupled receptors because "these membrane receptors are the targets for 60% to 70% of drugs," says Iwata. In the second paper, Huang3 shows how the membrane potential plays a critical role in protein unfolding and translocation across the outer and inner membranes in yeast mitochondria. In his review, F1000 member Eduardo Rial noted, "The authors show that the electrical potential across the mitochondrial inner membrane drives the unfolding of the protein that is being imported." While many re-searchers study protein folding, Matouschek's group focuses on unfolding, using isolated yeast mitochondria. Many mitochondrial proteins are synthesized in the cytosol with N-terminal targeting sequences and must then reach the right organelle. Matouschek says the current thinking is that acidic binding sites in an outer mitochondrial pore guide the positively charged targeting sequences to the inner membrane. Due to size constraints imposed by the translocation channels in the two membranes, the protein must unfold at the surface to reach the mitochondrial matrix. Using different-length targeting sequences and membrane potential modifiers, Matouschek shows that this unfolding is catalyzed from a distance when the membrane potential pulls at the charges in the targeting sequence. Matouschek uses the analogy of a tied shoe, explaining that force exerted at a taut shoelace's end passes through the shoelace to the knot, providing energy for unfolding. Once the polypeptide advances past the outer membrane and through the tight inner membrane pore, Matouschek notes, the ATP-driven mitochondrial Hsp70 motor kicks in to overcome friction and to "just rip the rest of the protein through." He adds that despite the presence of the ATP-driven Hsp70 motor, if the membrane potential is knocked out, no translocation occurs, no matter the length and positive charge of the targeting sequence. Says Matouschek, "If you don't have the membrane potential, you never get import. End of story." Susan Jenkins can be contacted at sjenkins@the-scientist.com. References 1. P. Mitchell, "Coupling of phosphorylation to electron and hydrogen transfer by a chemi-osmotic type of mechanism," Nature, 191:144-8, 1961. 2. M. Jormakka et al., "Molecular basis of proton motive force generation: Structure of formate dehydrogenase-N," Science, 295:1863-8, March 8, 2002. 3. S. Huang et al., "Protein unfolding by the mitochondrial membrane potential," Nature Structural Biology, 9:301-7, April 2002. 4. F.N. Magill, ed., The Nobel Prize Winners, vol. 3, Pasadena, Calif.: Salem Press, 1990, pp. 993-1001. 5. www.nobel.se/chemistry/laureates/1978/press.html ©2002, The Scientist Inc. Comment E-mail article

Form and Function Finally Prove Mitchell's Proton Motive Force

Form and Function Finally Prove Mitchell's Proton Motive Force

Perseverance, perspiration and a creative bent pay off | By Susan Jenkins

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