Extremophiles: They Love Living on the Edge

Image: Courtesy of NASA/Marshall Space Flight CenterImage: Courtesy of John Reeve FROZEN FLUFF: Using electron beams to drill holes, scientists recovered these extremophiles from Antarctic ice. Brent Christner (above, middle), a graduate student in John Reeve's lab at Ohio State University, collecting ice core samples in Antarctica. They thrive, not just survive. The microbes live in places such as the Antarctic, where temperatures reach -45°C in the summer; on the ocean floor, in constant 2°C waters; or in boiling hot springs. Aptly dubbed extremophiles, they have numerous scientists asking how, and what, they can learn from these remarkable organisms. The temperatures and conditions where extremophiles exist mimic environments in outer space, and some scientists are searching to see just what kind of connection, if any, exists between there and here. Other scientists are looking for more earthly connections; can these extremists improve life here on Earth, such as helping to lower cholesterol or improve laundry soap? New discoveries continue to occur. This year, Japanese investigators found a thermoacidophilic crenarchaeote, Sulfolobus sp. strain 7, in an 80°C hot spring with a pH between 2.5 and 3.1 Chinese scientists also reported isolating Marinospirillum alkaliphilum sp. Nov. from water in Inner Mongolia,2 where the pH is 9.5. A third team reported finding a bacterium with a protease that is "suitable for use in bleach-based detergents."3 Perhaps one of the most dramatic, and scientifically important roles extremophiles have played is in PCR. This DNA-amplifying technology--the basis of a several hundred million dollar a year enterprise--depends on the enzyme Taq polymerase. The enzyme was isolated from the bacterium Thermus aquaticus, discovered by University of Wisconsin professor of bacteriology Thomas Brock in 1966 (decades before PCR was developed) in hot springs in Yellowstone National Park.4 But it was the discovery of extrem-ophiles themselves, which occurred more than two decades ago, that changed how some scientists perceive life's parameters. "I think the real surprise is about our limited understanding of what the actual physical limits are for life," says senior scientist Edward DeLong, Monterey Bay Aquarium Research Institute in California. "Twenty five or 30 years ago, the upper temperature limit for life was [thought to be] near 60°C." EXTREME DISCOVERY It was Karl Stetter, professor of microbiology, University of Regensburg, Germany, who stirred interest in such organisms.5 Stetter found that some microbes live in extreme heat, calling the oceans' hydrothermal vents home. "[Karl] started to get pure cultures of organisms whose optimal growth temperature was 80°C," says DeLong, "and some of the isolates grew in boiling water. That challenged a lot of our concepts about the stability of the stuff that makes up life itself--macromolecules like proteins and DNA." DeLong notes that "microbial life is now known to exist at temperatures of at least 113°C, at pH values approaching zero, and at salinities found in saturated brines."6 When possible microfossils were found in an ancient Mars meteorite in the mid-1990s, scientists began wondering if there was any connection between extrem-ophiles here, and microfossils there. "You feel a bit differently about where you would look in the solar system where life might be hanging out," says Kenneth Nealson, a senior scientist at the Jet Propulsion Laboratory in Pasadena, Calif., and professor of earth sciences at the University of Southern California. "If you look at the nearby planets to which we have access, like Mars and the moon of Jupiter, Europa, the conditions which we believe are prevailing on those planets are conditions which we here on Earth would describe as extreme. So on Europa there is thought to be something like 4-km-thick ice and there could be water underneath." Says biochemistry professor Michael Danson of the UK's University of Bath, "By studying how organisms live and thrive in places like the Antarctic, if we can understand how those organisms operate, then we will have a good starting base by which to find life and study that life on ... Europa." Prokaryotes are the organisms that can function under these conditions. These "environmental tough guys," says Nealson, are the "Timex watches of life," meaning they do not die easily. Their rigid cell walls prevent them from engulfing prey. Instead, says Nealson, they survive as chemists. MYSTERY SURVIVORS Extremophiles do not seem to possess any outrageous measures that keep them alive, which is a bit disappointing to researchers. "There's no consistent or unique phenomenon," says microbiology professor John Reeve, Ohio State University. "That's probably the lesson that everyone has learned, which is not as exciting as you might want it. The biology is fairly standard. There isn't anything that you can immediately point to and say 'that's the reason it grows at 100°or zero degrees.' It's a combination of more stable proteins, more fluid or less fluid membranes ... extremes of what we already knew." At high temperatures, says Danson, the organisms' proteins and other large molecules might have "slightly" more interactions--more noncovalent forces at high temperatures and fewer at colder temperatures. "What an organism has to do is keep its molecules together," so more interactions would resist the forces that drive them apart. Photo: Courtesy of UGA Photographic Services Michael Adams The structural differences between proteins that remain stable at high temperatures and those that do not can be quite subtle, notes biochemistry professor Michael Adams, University of Georgia. A protein chemist studying two proteins, one stable at 100°C and one that falls apart at 25°C, would be "hard pressed to tell you which is which," he says. Adams adds that each protein has worked out its own solution to the problem of stabilization at high temperatures, eliminating the temptation to presume that only one stabilization mechanism exists. Moreover, says Danson, temperature does not affect the way the proteins are folded. He studied this phenomenon in the enzyme citrate synthase, which is the first enzyme in the Krebs cycle, the central pathway for all metabolism. In organisms grown in temperatures between 15° and 100°C, Danson found remarkable similarity. "What you can do is lay one structure over another and the basic fold is precisely the same--amazing conservation," he says. Adams was also surprised that the same folding pattern is used across that entire temperature range. Adams is currently working on pyrococcus furiosus, which was isolated in the mid-1980s from an oceanic volcanic vent off the Italian coast, and subsequently sequenced and found to have slightly more than 2,000 base pairs. "We're ... cloning all 2,000-plus genes, expressing them and obtaining the recombinant proteins, which we hand over to our crystallographic and NMR [nuclear magnetic resonance] collaborators for structural determination," he explains. The University of Georgia is one of nine centers established by the National Institutes of Health for such structural protein work. EXTREMOPHILES IN BUSINESS Extremophiles do more than challenge basic research questions and stimulate wonder about the possibility of extraterrestrial life. Their enzymes can be a boon to industrial processes, notes Reeve. To begin with, he says, enzymes can do "very precise chemistry; things a synthetic chemist would have a very difficult time doing." An extremophiles' enzymes can operate at the outrageous temperatures and pressures demanded by industrial processes. They can even present solutions to mundane tasks-like doing the laundry. Staff scientist Brian Jones at Genencor in Leiden, The Netherlands, has found bacteria of the genus Natrono in lakes in East Africa's Great Rift Valley. "You can think of it as being a caustic brine. We're talking about environments with pH 8 to pH 12. We're talking about lakes where there is so much salt that it crystallizes out so that you can walk across them," he says. Genencor, which screens these bacteria for useful enzymes, has found one that, when added to detergents, gives denim jeans that popular stonewashed look. Finding enzymes for use in cleaning products is one of the company's targeted areas. "Many of our applications for which we wish to use enzymes or products have to be particularly robust," says Jones. "A high-pH, watery environment with salts is probably Nature's nearest equivalent [to what goes on inside a washing machine]. That was one of the rationales behind looking for cleaning enzymes," he says. Researchers also search for these enzymes to improve industrial processes. One major void in making something as ordinary as high fructose corn syrup, a major food industry product, is the absence of enzymes that are stable at high temperatures, says Adams. "... In the past they [would] throw in huge amounts of very unstable enzymes. That makes the process work," he says. But that's not very efficient. "So if you can replace those with highly efficient, highly active enzymes, that seems worthwhile," Adams comments. Research scientist Peter Nichols, at the Commonwealth Scientific and Industrial Research Organization in Australia, says that investigators are looking at the industrial and related applications of psychrophiles, extremophiles found in the Arctic and Antarctic. "Very cold environments are known to increase the level of polyunsaturates in cell membranes. We have discovered bacteria that have those unsaturates," he says. Polyunsaturated fatty acids (PUFA), such as the omega-3 fatty acids that are considered valuable in lowering cholesterol, can be important additives to food, says Nichols. Fish, which eat these organisms, contain this oil. "With global fisheries under pressure and an increasing demand on omega-3 fatty acids for various nutriceutical purposes, new sources are needed," he says. But he cautions that while these acids have been found in microorganisms in the Antarctic,7 commercializing them on their home turf is virtually impossible. But maybe, Nichols says, scientists can transfer the PUFA genes into land plants that prefer higher thermostat readings. Harvey Black (hblack@chorus.net) is a freelance writer in Madison, Wis. References 1. T. Suzuki et al., "Sulfolobus tokodaii sp. nov. (f. Sulfolobus sp. strain 7), a new member of the genus Sulfolobus isolated from Beppu Hot Springs, Japan," Extremophiles, 6:39-44, February 2002. 2. W. Zhang et al., "Marinospirillum alkaliphilum sp. nov., a new alkaliphilic helical bacterium from Haoji soda lake in inner Mongolia autonomous region of China," Extremophiles, 6:33-7, February 2002. 3. K. Saeki et al., "A novel species of alkaliphilic Bacillus that produces an oxidatively stable alkaline serine protease," Extremophiles, 6:65-72, February 2002. 4. T. Brock, H. Freeze, "Thermus aquaticus gen. n. and sp. n., a nonsporulating extreme thermophile," Journal of Bacteriology, 98:289-97, 1969. 5. F. Fischer, et al., "Chemolithoautotrophic metabolism of anaerobic extremely thermophilic archaebacteria," Nature, 301:511-3, 1983. 6. E.F. DeLong, "Extreme genomes," online at www.genomebiology.com. 7. D. Nichols et al., "Developments with Antarctic microorganisms: culture collections, bioactivity screening, taxonomy, PUFA production and cold-adapted enzymes," Environmental Biotechnology, 10:240-6, 1999.

Extremophiles: They Love Living on the Edge

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Image: Courtesy of NASA/Marshall Space Flight Center

Image: Courtesy of John Reeve

FROZEN FLUFF: Using electron beams to drill holes, scientists recovered these extremophiles from Antarctic ice. Brent Christner (above, middle), a graduate student in John Reeve's lab at Ohio State University, collecting ice core samples in Antarctica.