Group photo. Members of the Biotechnology and Genetics of Thermophilic Bacteria group. Standing from left: Leticia Torres, Eloy Ferreras, Marcos Almendros, José Berenguer, Carlos Bricio, Esther Sanchez, Laura Alvarez, Jorge Perez. Seated: Aurelio Hidalgo, Maria Luisa del Pozo, Alba Blesa. Center for Molecular Biology «Severo Ochoa» CSIC-Autonomous University of Madrid
Over the last 20 years, thermophilic organisms have become very relevant biological models in different fields of Biosciences. One such aspect is the use that has been made of the intrinsic stability of its components to develop biotechnological applications of enormous repercussion, such as the DNA amplification process, which has meant an extraordinary leap in our ability to detect genetic information in tests. diagnostic, forensic, and in all areas of Biology. In addition, given the association between thermal stability and resistance to organic solvents and detergents, there is great interest in the industry for the use of these enzymes in biocatalysis processes. A second relevant aspect is given by the greater facility for crystallization of proteins and large biological complexes of thermophiles. Thanks to this, the first high-resolution structures obtained from ribosomes, the machines that manufacture proteins, or from the so-called respiratory complex I, a fundamental piece for respiration, were obtained from thermophilic bacteria. Therefore, thermophilic organisms also constitute excellent models in Structural Biology. Finally, the thermophilic organisms that grow at higher temperatures are the most similar to the first living beings that inhabited this planet. Although this aspect is still controversial, most of the genetic comparisons between organisms point to thermophilic organisms as the oldest in evolution, their analysis constituting a way of traveling through time to discover what the first inhabitants of the earth were like.
Despite this biological and applied interest, the use of thermophilic organisms as a model is restricted by the difficulty that their cultivation presents for a laboratory. Much effort has been used by various laboratories in the world to "domesticate" some of these organisms. Despite this, only reasonable success has been achieved with archaea Sulfolobus spp y Thermococcus spp, and the bacteria Thermus thermophilus.
Our research group has been one of those that has participated in the domestication and adaptation to the laboratory of Thermus thermophilus, and today it constitutes our main work model. Unlike other extreme thermophilic bacteria, many isolates from T. thermophilus they grow rapidly in the laboratory, doubling their population at 70ºC every 45 min in liquid media, and forming plate colonies in 24 hours. Furthermore, the availability of a highly efficient natural competition apparatus has allowed us to develop a complete set of genetic tools that gives access to its physiological and functional analysis, and even to its use as a cellular factory for the production of proteins.
Using this model, in our laboratory we follow two parallel lines of research. On the one hand, we study the denitrification process present in some strains of Thermus spp and its regulation, and the way in which this capacity is transferred horizontally, and on the other, we develop biotechnological applications derived from the organism or its enzymes.
Denitrification is known as a process in which nitrogen oxides are used instead of oxygen to burn nutrients and obtain energy, while eliminating nitrate from the medium. Our group has described and characterized a set of reductases necessary to breathe nitrate (Nar), nitrite (Nir) and nitric oxide (Nor) at high temperature, and we have discovered that the genes that encode them are integrated in a region of the genome that is easily transferable to other strains of the same species. Among the most interesting biochemical and functional aspects that we have described is the fact that Nar from T. thermophilus, in addition to reducing nitrate, is capable of acting as an electron carrier towards Nir and Nor, something that in non-thermophilic organisms is carried out by the respiratory complex III, which Nar replaces in this role. Other important aspects have been described for Nor, which contains an additional subunit of unknown function and has several proton entry pathways from the cytoplasm. At a more genetic level, we are intrigued by the mechanism of DNA transfer by direct cell-cell contact, since it does not resemble any of those described so far. It is interesting to note that during its study compared to the natural transformation system we have discovered the existence of a protein similar to the human Argonaut that protects cells through a unique DNA-DNA interference system, the first of its kind that has been described, of the possible harmful action of genes of unknown origin acquired from the environment.
In a more applied line, our efforts have focused mainly on two aspects. On the one hand, we have developed a procedure that allows the selection of thermostable forms of enzymes and proteins from non-thermophilic organisms by folding interference. This technique consists of expressing in our model bacteria at high temperature fusions between the protein to be stabilized and a protein that confers a detectable property, such as resistance to an antibiotic. Normally, the unstable protein will misfold under these conditions and interfere with the folding of the control, giving rise to sensitive bacteria. In contrast, thermostable variants will fold well, will not interfere, and will generate resistant clones. In this way we have stabilized from therapeutic proteins to enzymes useful in biocatalysis. In the immediate future and through EU and MINECO projects, we will develop high-capacity screening versions of this method, using thermostable variants of fluorescent proteins that we have developed. On the other hand, we have used enzymes from different strains for use in biocatalysis processes.
Representative bibliography
Cava F, Hidalgo A, Berenguer J. (2019). A Live Thermus thermophilus Cava F, Hidalgo A, Berenguer J.
Cava F, Hidalgo A, Berenguer J. Cava F, Hidalgo A, Berenguer J. Thermus thermophilus Cava F, Hidalgo A, Berenguer J.
Cava F, Hidalgo A, Berenguer J. Cava F, Hidalgo A, Berenguer J.
Cava F, Hidalgo A, Berenguer J. Cava F, Hidalgo A, Berenguer J. Thermus thermophilusCava F, Hidalgo A, Berenguer J.
Cava F, Hidalgo A, Berenguer J. Cava F, Hidalgo A, Berenguer J. Thermus thermophilus Cava F, Hidalgo A, Berenguer J.
Cava F, Hidalgo A, Berenguer J. (2008). A cytochrome c containing nitrate reductase plays a role in electron transport for denitrification in Thermus thermophilus (2008). A cytochrome c containing nitrate reductase plays a role in electron transport for denitrification in
(2008). A cytochrome c containing nitrate reductase plays a role in electron transport for denitrification in (2008). A cytochrome c containing nitrate reductase plays a role in electron transport for denitrification in:(2008). A cytochrome c containing nitrate reductase plays a role in electron transport for denitrification in
(2008). A cytochrome c containing nitrate reductase plays a role in electron transport for denitrification in (2008). A cytochrome c containing nitrate reductase plays a role in electron transport for denitrification in Thermus thermophilus(2008). A cytochrome c containing nitrate reductase plays a role in electron transport for denitrification in
(2008). A cytochrome c containing nitrate reductase plays a role in electron transport for denitrification in (2008). A cytochrome c containing nitrate reductase plays a role in electron transport for denitrification in:(2008). A cytochrome c containing nitrate reductase plays a role in electron transport for denitrification in
Swarts DC, Jore MM, Westra ER, Zhu Y, Janssen JH, Snijders AP, Wang Y, Patel DJ, Berenguer J, Brouns SJ, Van Der Oost J. Swarts DC, Jore MM, Westra ER, Zhu Y, Janssen JH, Snijders AP, Wang Y, Patel DJ, Berenguer J, Brouns SJ, Van Der Oost J.:Swarts DC, Jore MM, Westra ER, Zhu Y, Janssen JH, Snijders AP, Wang Y, Patel DJ, Berenguer J, Brouns SJ, Van Der Oost J.
Swarts DC, Jore MM, Westra ER, Zhu Y, Janssen JH, Snijders AP, Wang Y, Patel DJ, Berenguer J, Brouns SJ, Van Der Oost J. Swarts DC, Jore MM, Westra ER, Zhu Y, Janssen JH, Snijders AP, Wang Y, Patel DJ, Berenguer J, Brouns SJ, Van Der Oost J. 4:Swarts DC, Jore MM, Westra ER, Zhu Y, Janssen JH, Snijders AP, Wang Y, Patel DJ, Berenguer J, Brouns SJ, Van Der Oost J.
Swarts DC, Jore MM, Westra ER, Zhu Y, Janssen JH, Snijders AP, Wang Y, Patel DJ, Berenguer J, Brouns SJ, Van Der Oost J. Swarts DC, Jore MM, Westra ER, Zhu Y, Janssen JH, Snijders AP, Wang Y, Patel DJ, Berenguer J, Brouns SJ, Van Der Oost J. Thermus thermophilus Swarts DC, Jore MM, Westra ER, Zhu Y, Janssen JH, Snijders AP, Wang Y, Patel DJ, Berenguer J, Brouns SJ, Van Der Oost J.:Swarts DC, Jore MM, Westra ER, Zhu Y, Janssen JH, Snijders AP, Wang Y, Patel DJ, Berenguer J, Brouns SJ, Van Der Oost J.
Swarts DC, Jore MM, Westra ER, Zhu Y, Janssen JH, Snijders AP, Wang Y, Patel DJ, Berenguer J, Brouns SJ, Van Der Oost J. Swarts DC, Jore MM, Westra ER, Zhu Y, Janssen JH, Snijders AP, Wang Y, Patel DJ, Berenguer J, Brouns SJ, Van Der Oost J. Thermus thermophilus Swarts DC, Jore MM, Westra ER, Zhu Y, Janssen JH, Snijders AP, Wang Y, Patel DJ, Berenguer J, Brouns SJ, Van Der Oost J.Swarts DC, Jore MM, Westra ER, Zhu Y, Janssen JH, Snijders AP, Wang Y, Patel DJ, Berenguer J, Brouns SJ, Van Der Oost J.Swarts DC, Jore MM, Westra ER, Zhu Y, Janssen JH, Snijders AP, Wang Y, Patel DJ, Berenguer J, Brouns SJ, Van Der Oost J. 11:101.
101. 101. Thermus thermophilus 101.
101. 101.
101. 101. Thermus thermophilus101.