NEWS OF THE WEEK
ELECTRONSTARVED ENZYME ENZYME CATALYSIS: Cytochrome c
oxidase model mimics natural electron-limited conditions
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NEW MODEL of the active site of a key enzyme in cellular respiration allows scientists to study the enzyme under conditions where the flow of electrons is the limiting step, as it is in the natural enzyme (Science 2007,315,1565). Cytochrome c oxidase catalyzes the four-electron reduction of 0 2 to H 2 0 during the final stage of respiration. This reduction must happen without releasing partially reduced oxygen species, which are toxic. How the enzyme accomplishes this is poorly understood, because scientists haven't been able to study the enzyme under electron-limited conditions that might be expected to lead to partial reductions. "The enzyme is always starved for electrons, something few people seem to recognize," says chemistry professor James P. Collman of Stanford University. Collman worked with graduate student Neal K. Devaraj, research associate Richard A. Decréau, and coworkers to build a biologically relevant model system that includes three redox sites: a myoglobin-like heme, copper suspended among three imidazoles about 5 Â above the heme, and a phenol group covalently attached to one of the copper-ligating imidazoles. In earlier models, Collman and coworkers adsorbed the catalysts on graphite electrodes, so the electron transfer was very rapid. Now, in collaboration with Stanford chemistry professor Christopher E. D. Chidsey, they slow the electron transfer by fastening the catalyst covalently onto a gold electrode modified
with a self-assembled monolayer. The anchored model system answers some elusive questions about cytochrome c oxidase's mechanism, Collman says. "Biologists have usually studied cytochrome c oxidase by loading it with electrons and allowing it to turn over one time. It's really difficult for biologists to study cytochrome c oxidase under steady-state 1 ^ 0 conditions and see what happens right at the active site," he says. For example, biologists have suspected that all three of the natural M I M I C A model of cytochrome c oxienzyme's redox sites were necessary dase reproduces the key components of for preventing the release of partially the enzyme active site, including iron, reduced oxygen species, but it's been copper, and phenol redox sites. Attaching difficult to demonstrate. "By protectthe synthetic system to self-assembled ing in turn each of the redox-active monolayers gates the flow of electrons. substituents, we're able to show that to get the four-electron reduction that cytochrome c oxidase must do in order to prevent the formation of these toxic partially reduced oxygen species, all three substituents must be present at once," Collman explains. Constructing the model has been a longterm effort, Collman says, culminating with the ad*^.~ dition of the crucial phenol, which mimics a key tyrosine residue in the natural enzyme. Constructing the model requires a 32-step convergent synthesis. The model doesn't mimic everything the natural enzyme does, Collman points out. With a fully oxidized enzyme, the first electron goes to a peripheral redox site outside of the active site. When a second electron is Crystal structure added, a conformational change causes both electrons of cytochrome c to jump to the active site. In this way, the enzyme avoids oxidase active site forming partially reduced oxygen species. "We have not (red is Fe, green is been able to imitate that," Collman says. "That's someCM, black is N, gray thing we would like to do."-CELIA ARNAUD is C, blue is O).
VENTER'S ADVENTURE Survey of marine microbes reveals a wealth of genetic diversity Some 7.7 million DNA sequences containing 6.3 billion base pairs—along with 6.1 million proteins, including thousands of protein kinases and other enzymes—make up the treasure trove of data extracted from ocean-dwelling bacteria and viruses during the two-year Sorcerer II Global Ocean Sampling Expedition. Research papers and associated articles reporting these findings were published on March 13 in the online journal PLoS Biology (collec tions.plos.org/plosbiology/gos-2007.php). This vast amount of sequence data, gathered and processed by researchers at the J. Craig Venter Institute, in Rock-
ville, Md., is expected to lead to better understanding of key biological processes. For example, information on families of proteins gleaned from the data could help clarify the ocean's role in global climate and eventually lead to new antibiotics and other medicines, technologies for alternative energy production, and methods for industrial processing. The new data, which are being freely shared via public databases on the Web, is just the beginning. Additional surveys are planned for sampling at different locations and ocean depths. "Given the findings, it's clear that we've
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only begun to scratch the surface of understanding the microbial world around us," says expedition leader J. Craig Venter, the institute's founder and chairman. Access to the information on genetic diversity is "wonderful and extremely valuable," comments Per Falholt, chief scientific officer at Novozymes, a global leader in developing enzymes for industrial applications. "But it's just the start of another mountain we need to climb." The volume of data presents a huge task for bioinformatics experts to sort through and make links between protein functions and potential applications, he says.-STEVE RITTER
