Overview

Brief Summary

Thermus aquaticus is a thermophilic gram-negative bacterium that has played a key role in the modern revolution in genetic research, genetic engineering, and biotechnology (see below). Thermophilic bacteria are bacteria that thrive at very high temperatures, often above 45° C (113° F). Thermus aquaticus was originally isolated from a number of hot springs in Yellowstone National Park and a hot spring in California (U.S.A.) and was subsequently isolated from hot springs in other parts of the world and even from artificial hot water environments such as hot tap water (Brock and Freeze 1969; Brock and Boylen 1973). Previously, microbiologists had enriched for thermophilic bacteria at 55° C, but the discoverer of T. aquaticus, Thomas Brock, found that many thermophiles in his studies of microbial ecology would not be easily detected at such a "low" temperature since they require temperatures above 70° C to flourish. As Brock has noted, his discovery provides an excellent illustration of the often unpredictable value (both academic and applied) of basic research (Brock 1997).

In the 1980s, a method known as the polymerase chain reaction (PCR) was developed to generate many copies of targeted segments of DNA from very tiny samples (Mullis and Faloona 1987; Arnheim and Erlich 1992; visit this site for a visual explanation of the principle of PCR) . This technique includes repeated cycles of melting apart of the two strands of each double-stranded DNA molecule (typically at 92° to 95° C) alternating with the extension of new complementary strands to create additional copies. To be practical, this method requires the use of a DNA polymerase that is not destroyed by this heating (DNA polymerases are enzymes that play a key role in DNA synthesis within cells; see Pavlov et al. 2004 for an overview of DNA polymerases). Fortunately, evolution has provided such polymerases in bacteria adapted to live at very high temperatures (e.g., in hot springs). Chien et al. (1976) had already purified a stable DNA polymerase from T. aquaticus with a temperature optimum of 80° C, which proved to serve very well for the automation of PCR.

The use of DNA polymerases from T. aquaticus and other thermophiles in PCR and related applications, such as DNA sequencing, has revolutionized biotechnology. The humble T. aquaticus enormously expanded what questions could be practically addressed in fields ranging from biomedical science ("what is the genetic basis for disease X and does this patient have this disorder?") to animal behavior ("were all the young in this bluebird nest actually sired by the mother's apparent mate?") to conservation ("is this whale meat being sold truly from the species the seller claims?") to forensics ("can this accused criminal possibly have left the DNA evidence found at the crime scene?") and beyond.

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Comprehensive Description

Thermus aquaticus is a thermophilic gram-negative bacterium that has played a key role in the modern revolution in genetic research, genetic engineering, and biotechnology (see below). Thermophilic bacteria are bacteria that thrive at very high temperatures, often above 45° C (113° F). Thermus aquaticus was originally isolated from a number of hot springs in Yellowstone National Park and a hot spring in California (U.S.A.) and was subsequently isolated from hot springs in other parts of the world and even from artificial hot water environments such as hot tap water (Brock and Freeze 1969; Brock and Boylen 1973). Previously, microbiologists had enriched for thermophilic bacteria at 55° C, but the discoverer of T. aquaticus, Thomas Brock, found that many thermophiles in his studies of microbial ecology would not be easily detected at such a "low" temperature since they require temperatures above 70° C to flourish. As Brock has noted, his discovery provides an excellent illustration of the often unpredictable value (both academic and applied) of basic research (Brock 1997).

In the 1980s, a method known as the polymerase chain reaction (PCR) was developed to generate many copies of targeted segments of DNA from very tiny samples (Mullis and Faloona 1987; Arnheim and Erlich 1992; visit this site for a visual explanation of the principle of PCR) . This technique includes repeated cycles of melting apart of the two strands of each double-stranded DNA molecule (typically at 92° to 95° C) alternating with the extension of new complementary strands to create additional copies. To be practical, this method requires the use of a DNA polymerase that is not destroyed by this heating (DNA polymerases are enzymes that play a key role in DNA synthesis within cells; see Pavlov et al. 2004 for an overview of DNA polymerases). Fortunately, evolution has provided such polymerases in bacteria adapted to live at very high temperatures (e.g., in hot springs). Chien et al. (1976) had already purified a stable DNA polymerase from T. aquaticus with a temperature optimum of 80° C, which proved to serve very well for the automation of PCR. The use of DNA polymerases from T. aquaticus and other thermophiles in PCR and related applications, such as DNA sequencing, has revolutionized biotechnology. The humble T. aquaticus enormously expanded what questions could be practically addressed in fields ranging from biomedical science ("what is the genetic basis for disease X and does this patient have this disorder?") to animal behavior ("were all the young in this bluebird nest actually sired by the mother's apparent mate?") to conservation ("is this whale meat being sold truly from the species the seller claims?") to forensics ("can this accused criminal possibly have left the DNA evidence found at the crime scene?") and beyond.

Gibbs et al. (2009) studied the phylogeny of Thermus isolates and found they fell into 8 clades. They then isolated the DNA polymerase I genes from 22 representatives and cloned the 8 most diverse genes (to represent the 8 clades) into an expression vector. They were able to purify the protein from six of these clones and examined their biochemical characteristics and suitability for PCR. They found that none was as thermostable as commercially available Taq polymerase and all had error-frequencies similar to those of Taq polymerase. They concluded that the initial selection of T. aquaticus for DNA polymerase purification was a fortuitous choice, although they suggest that simple mutagenesis procedures on other Thermus-derived polymerases should provide comparable thermostability for the PCR reaction.

In addition to its DNA polymerase, a range of other thermostable enzymes have been isolated from T. aquaticus for use in high temperature molecular biology applications. Largely as a result of the example of T. aquaticus, which has generated enormous profits for the biotechnology industry but not for the national park in which it was discovered (nor for U.S. taxpayers), the National Park Service now may require researchers working in national parks to sign "benefits sharing" agreements requiring that profits that may be derived at a later point in time from work done in the park be shared the park.

Brock and Edwards (1970) reported on the fine structure of Thermus aquaticus.

Degryse et al. (1978) and Degryse and Glansdorff (1981) studied the metabolism of Thermus aquaticus. These bacteria are obligately aerobic and chemoheterotrophic (Degryse et al. 1978).

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Ecology

Habitat

Thermus aquaticus was originally isolated from a number of hot springs in Yellowstone National Park and a hot spring in California (U.S.A.) and was subsequently isolated from hot springs in other parts of the world and even from artificial hot water environments such as hot tap water (Brock and Freeze 1969; Brock and Boylen 1973). Previously, microbiologists had enriched for thermophilic bacteria (i.e, bacteria that thrive at high temperatures) at 55° C, but the discoverer of T. aquaticus, Thomas Brock, found that many thermophiles in his studies of microbial ecology would not be easily detected at such a "low" temperature since they require temperatures above 70° C to flourish.

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Evolution and Systematics

Systematics or Phylogenetics

Gibbs et al. (2009) studied the phylogeny of Thermus isolates and found they fell into 8 clades. They then isolated the DNA polymerase I genes from 22 representatives and cloned the 8 most diverse genes (to represent the 8 clades) into an expression vector. They were able to purify the protein from six of these clones and examined their biochemical characteristics and suitability for PCR. They found that none was as thermostable as commercially available Taq polymerase and all had error-frequencies similar to those of Taq polymerase. They concluded that the initial selection of T. aquaticus for DNA polymerase purification was a fortuitous choice, although they suggest that simple mutagenesis procedures on other Thermus-derived polymerases should provide comparable thermostability for the PCR reaction.

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Physiology and Cell Biology

Physiology

Degryse et al. (1978) and Degryse and Glansdorff (1981) studied the metabolism of Thermus aquaticus. These bacteria are obligately aerobic and chemoheterotrophic (Degryse et al. 1978).

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Relevance to Humans and Ecosystems

Benefits

The use of DNA polymerases from T. aquaticus and other thermophiles in the polymerase chain reaction (PCR) and related applications, such as DNA sequencing, has revolutionized biotechnology. The humble T. aquaticus enormously expanded what questions could be practically addressed in fields ranging from biomedical science ("what is the genetic basis for disease X and does this patient have this disorder?") to animal behavior ("were all the young in this bluebird nest actually sired by the mother's apparent mate?") to conservation ("is this whale meat being sold truly from the species the seller claims?") to forensics ("can this accused criminal possibly have left the DNA evidence found at the crime scene?") and beyond.

In addition to its DNA polymerase, a range of other thermostable enzymes have been isolated from T. aquaticus for use in high temperature molecular biology applications. Largely as a result of the example of T. aquaticus, which has generated enormous profits for the biotechnology industry but not for the national park in which it was discovered (nor for U.S. taxpayers), the National Park Service now may require researchers working in national parks to sign "benefits sharing" agreements requiring that profits that may be derived at a later point in time from work done in the park be shared with the park.

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Wikipedia

Thermus aquaticus

Thermus aquaticus is a species of bacterium that can tolerate high temperatures, one of several thermophilic bacteria that belong to the Deinococcus-Thermus group. It is the source of the heat-resistant enzyme Taq DNA polymerase, one of the most important enzymes in molecular biology because of its use in the polymerase chain reaction (PCR) DNA amplification technique.

History[edit]

When studies of biological organisms in hot springs began in the 1960s, scientists thought that the life of thermophilic bacteria could not be sustained in temperatures above about 55 °C (131 °F).[1] Soon, however, it was discovered that many bacteria in different springs not only survived, but also thrived in higher temperatures. In 1969, Thomas D. Brock and Hudson Freeze of Indiana University reported a new species of thermophilic bacterium which they named Thermus aquaticus.[2] The bacterium was first discovered in the Lower Geyser Basin of Yellowstone National Park, near the major geysers Great Fountain Geyser and White Dome Geyser,[3] and has since been found in similar thermal habitats around the world.

Biology[edit]

It thrives at 70°C (158°F), but can survive at temperatures of 50°C to 80°C (122°F to 176°F). This bacterium is a chemotroph — it performs chemosynthesis to obtain food. However, since its range of temperature overlaps somewhat with that of the photosynthetic cyanobacteria that share its ideal environment, it is sometimes found living jointly with its neighbors, obtaining energy for growth from their photosynthesis.

Morphology[edit]

Thermus aquaticus is generally of cylindrical shape with a diameter of 0.5 μm to 0.8 μm. The shorter rod shape has a length of 5 μm to 10 μm. The longer filament shape has a length that varies greatly and in some cases exceeds 200 μm. The rod-shaped bacteria have a tendency to aggregate. Associations of several individuals can lead to the formation of spherical bodies 10 μm to 20 μm in diameter, also called rotund bodies.[2][4]

Enzymes from T. aquaticus[edit]

T. aquaticus has become famous as a source of thermostable enzymes, particularly the Taq DNA polymerase, as described below.

Aldolase
Studies of this extreme thermophilic bacterium that could be grown in cell culture was initially centered on attempts to understand how protein enzymes (which normally inactivate at high temperature) can function at high temperature in thermophiles. In 1970, Freeze and Brock published an article describing a thermostable aldolase enzyme from T. aquaticus.[5]
RNA polymerase enzyme isolated from T. aquaticus in 1974 was a DNA-dependent RNA polymerase,[6] used in the process of transcription.
Taq I restriction enzyme
For more details on this topic, see TaqI.
Most molecular biologists probably became aware of T. aquaticus in the late 1970s or early 1980s because of the isolation of useful restriction endonucleases from this organism.[7] Use of the term Taq to refer to Thermus aquaticus arose at this time from the convention of giving restriction enzymes sr".[8] In 1993, Dr. Mullis was awarded the Nobel Prize for his work with PCR.
Other enzymes
The high optimum temperature for T. aquaticus allows researchers to study reactions under conditions for which other enzymes lose activity. Other enzymes isolated from this organism include DNA ligase, alkaline phosphatase, NADH oxidase, isocitrate dehydrogenase, amylomaltase, and fructose 1,6-disphosphate-dependent L-lactate dehydrogenase.

Controversy[edit]

The commercial use of enzymes from T. aquaticus has not been without controversy. After Dr. Brock's studies, samples of the organism were deposited in the American Type Culture Collection, a public repository. Other scientists, including those at Cetus, obtained it from there. As the commercial potential of Taq polymerase became apparent in the 1990s,[9] the National Park Service labeled its use as the "Great Taq Rip-off".[10] Researchers working in National Parks are now required to sign "benefits sharing" agreements that would send a portion of later profits back to the Park Service.

See also[edit]

References[edit]

  1. ^ Thomas Brock's essay "Life at High Temperatures", available at[dead link]
  2. ^ a b Brock TD and Freeze H (1969). "Thermus aquaticus, a Nonsporulating Extreme Thermophile". J. Bact. 98 (1): 289–97. PMC 249935. PMID 5781580. 
  3. ^ Bryan, T. Scott (2008). Geysers of Yellowstone, The (4th ed.). University Press of Colorado. ISBN 978-0-87081-924-7. 
  4. ^ Brock TD and Edwards MR (1970). "Fine Structure of Thermus aquaticus, an Extreme Thermophile". J. Bact. 104 (1): 509–517. PMC 248237. PMID 5473907. 
  5. ^ Freeze H and Brock TD (1970). "Thermostable Aldolase from Thermus aquaticus". J. Bact. 101 (2): 541–50. PMC 284939. PMID 4984076. 
  6. ^ Air GM and Harris JI (1974). "DNA-Dependent RNA Polymerase From the Thermophilic Bacterium Thermus aquaticus". FEBS Letters 38 (3): 277–281. doi:10.1016/0014-5793(74)80072-4. PMID 4604362. 
  7. ^ Sato, S (February 1978). "A single cleavage of Simian virus 40 (SV40) DNA by a site specific endonuclease from Thermus aquaticus, Taq I". J. Biochem (Tokyo) 83 (2): 633–5. PMID 204628. 
  8. ^ Guyer RL, Koshland DE (December 1989). "The Molecule of the Year". Science 246 (4937): 1543–6. doi:10.1126/science.2688087. PMID 2688087. 
  9. ^ Fore J, Wiechers IR, Cook-Deegan R (2006). "The effects of business practices, licensing, and intellectual property on development and dissemination of the polymerase chain reaction: case study". J Biomed Discov Collab 1: 7. doi:10.1186/1747-5333-1-7. PMC 1523369. PMID 16817955.  — Detailed history of Cetus and the commercial aspects of PCR.
  10. ^ Robbins J (28 November 2006). "The Search for Private Profit in the Nation's Public Parks". The New York Times. 

Further reading[edit]

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