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Analysis of the T. thermophilus HB27 gen- ome sequence reveals a variety of interesting pathways and enzymes, including metabolic pathways for vitamin B12 and carotenoid biosynthesis, as well as various proteases, glycoside hydrolases, phosphatases, oxidore- ductases and DNA-modifying enzymes. Certain T. thermophilus enzymes are inter- esting candidates for biotechnological app- lications 5 , such as molecular biological techniques for research and diagnostic use (DNA and RNA processing enzymes), in the starch industry (starch-converting enzymes), or in the food, detergent or leather industries (proteases). Turning interesting enzymes identified from genome information into marketable enzyme is a long and arduous process, however. Success will depend not only on the catalytic and stability properties of a particular enzyme of interest but also on its performance under the often-stringent conditions of existing or newly developed industrial processes. One of the advantages of using ther- mophilic eubacteria rather than archaea as a source of industrial enzyme-encoding seq- uences is that eubacterial genes are often more readily expressed in Escherichia coli or other standard bacterial cloning hosts than archaeal genes. In fact, initial expression experiments often work with a recombinant genes’ own expression signals, which can greatly simplify cloning and expression screening procedures. Efficient expression in standard host bacteria is of further benefit when embarking on large-scale production of an enzyme of interest. Even with these advantages, however, sev- eral potential obstacles may also complicate recombinant expression of T. thermophilus proteins. The first problem in obtaining high- level expression may be the unusual composi- tion of T. thermophilus genes. The genus Thermus belongs to a deeply branching line- age in the bacterial evolutionary tree, with DNA with a high (69.4 %) G+C content. Although more practical experience is needed to show how well heterologous expression of T. thermophilus genes of interest work in stan- dard expression systems, several T. aquaticus and T. thermophilus genes have already been successfully expressed in E. coli. And even if high-level expression of a particularly inter- esting open reading frame (ORF) were to fail, resynthesis of the entire gene, taking into account the codon usage of the host organ- ism, could provide a solution. A second problem facing recombinant expression is folding. It is not clear whether recombinant expression in a mesophilic bac- terium will deleteriously affect the folding of 524 VOLUME 22 NUMBER 5 MAY 2004 NATURE BIOTECHNOLOGY The genome sequences of at least 14 extre- mely thermophilic microorganisms (with growth optima above 70 °C) and hyperther- mophilic microorganisms (optimum growth temperature at least 80 °C) have now been determined. The latest to be completed— reported in this issue by Henne et al. 1 —is the genome of Thermus thermophilus, an aerobic, obligately heterotrophic eubacterium capable of growth at temperatures up to ∼85 °C. T. thermophilus is one of only four extremely thermophilic eubacteria sequenced to date. With the genome sequence in hand and its relative amenability to genetic manipulation, T. thermophilus is now positioned as an inter- esting model organism for extremely ther- mophilic eubacteria. In the longer term, the availability of its genome sequence, which consists of a 1.9-Mbp chromosome and a 0.23-Mbp megaplasmid, should facilitate the identification of novel proteins and pathways of potential biotechnological interest. A survey of prokaryotic genome data avail- able over the web 2 shows that, thus far, many more eubacterial genomes (140) than arch- aeal genomes (18) have been sequenced (a ratio of about 8:1) 2 . Of the 14 genomes of extreme thermophiles and hyperthermo- philes, 10 are from archaea and only 4 from eubacteria. The three eubacteria previously sequenced—the chemolithoautotrophic hyp- erthermophile Aquifex aeolicus, the chemo- organotrophic hyperthermophile Thermo- toga maritima and the chemoorganotrophic extreme thermophile Thermoanaerobacter tengcongensis—have now been joined by Thermus thermophilus 1 . The abundance of genomes from archaea reflects the high repre- sentation of these microorganisms in extreme environments as compared with eubacteria. Studies on the biochemical, biophysical and structural characterization of enzymes from extreme thermophiles (particularly hyperthermophilic bacteria and archaea, such as Thermotoga or Pyrococcus sp.) have led to a wealth of information on thermostable bio- catalysts and other basic aspects of protein stabilization 3,4 . Because of their inherently robust nature, their insensitivity toward denaturing conditions (not only heat, but also chemical denaturants and organic solvents) and their reduced conformational flexibility at ambient temperature, proteins from extreme thermophiles are often the preferred choices as substrates for X-ray crystallogra- phy studies or templates for protein engineer- ing. The above properties are also desirable for biotechnological applications, where not are only biocatalytic characteristics (e.g., reac- tion specificity or conversion rates) impor- tant, but also ease of handling, shelf-life and stability may be critical. Tth (T. thermophilus) and Taq (Thermus aquaticus) DNA polymerases are already familiar to many molecular biologists as com- mercialized enzymes routinely employed in nucleic acid amplification procedures (e.g., polymerase chain reaction (PCR) amplifica- tion and reverse transcriptase (RT)-PCR). But with the T. thermophilus genome now in hand, we can expect to uncover much more about the unique and exotic enzymatic machinery of this exceptional organism. Wolfgang Liebl is at the Institut für Mikrobiologie und Genetik at the Georg- August-Universität, Grisebachstrasse 8, D-37077 Göttingen, Germany. e-mail: wliebl@gwdg.de Genomics taken to the extreme Wolfgang Liebl The genome sequence of Thermus thermophilus may provide new insights into heat-tolerant enzymes and metabolic pathways of biotechnological potential. © 2004 Nature Publishing Group http://www.nature.com/naturebiotechnology