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Saugstad, Jens; Frøland, Stig S.; Madsen, Kjell; Matlary, Janne Haaland; Sirevåg, Reidun & Preus, Inger Julie Nygaard
(2018).
Hva mener rektorene?
Aftenposten (morgenutg. : trykt utg.).
ISSN 0804-3116.
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Saugstad, Jens; Frøland, Stig S.; Madsen, Kjell; Matlary, Janne Haaland; Sirevåg, Reidun & Preus, Inger Julie Nygaard
(2018).
Orwelliansk nytale fra SAIH.
Aftenposten (morgenutg. : trykt utg.).
ISSN 0804-3116.
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Saugstad, Jens; Frøland, Stig S.; Madsen, Kjell; Matlary, Janne Haaland; Sirevåg, Reidun & Preus, Inger Julie Nygaard
(2018).
Avkolonisering og akademisk frihet.
Aftenposten (morgenutg. : trykt utg.).
ISSN 0804-3116.
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Saugstad, Jens; Frøland, Stig Sophus; Madsen, Kjell; Matlary, Janne Haaland; Sirevåg, Reidun & Preus, Inger Julie Nygaard
(2018).
Avkoloniseringsdebatten truer universitetene.
Aftenposten (morgenutg. : trykt utg.).
ISSN 0804-3116.
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Sirevåg, Reidun
(2018).
Diktatur kan være nyttig.
Aftenposten (morgenutg. : trykt utg.).
ISSN 0804-3116.
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Sirevåg, Reidun
(2018).
Et bioteknologisk eventyr med store politiske følger.
Aftenposten (morgenutg. : trykt utg.).
ISSN 0804-3116.
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Helsvig, Kim Gunnar; Sirevåg, Reidun & Lærum, Ole Didrik
(2006).
Norwegian Academy of Science and Letters. From National Policy to Global Science.
Allea Biennial Yearbook.
p. 121–133.
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Bjørk, Alexandra; Dalhus, Bjørn; Mantzilas, Dimitris; Eijsink, Vincent & Sirevåg, Reidun
(2004).
Stabilization of a tetrameric malate dehydrogenase.
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Bjørk, Alexandra; Dalhus, Bjørn; Mantzilas, Dimitrios; Eijsink, Vincent G. H. & Sirevåg, Reidun
(2004).
Stabilization of a tetrameric malate dehydrogenase.
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Bjørk, Alexandra; Mantzilas, Dimitris; Sirevåg, Reidun & Eijsink, Vincent
(2003).
Stabilization of tetrameric malate dehydrogenase from Chloroflexus aurantiacus by engineering of an ionic network in the dimer-dimer interface.
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Dalhus, Bjørn; Bjørk, Alexandra; Eijsink, Vincent G. H. & Sirevåg, Reidun
(2003).
Structural basis for protein thermostability; analysis of interactions within a protein oligomer and engeneering towards higher thermostability.
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Bjørk, Alexandra; Dalhus, Bjørn; Mantzilas, Dimitrios; Eijsink, Vincent G. H. & Sirevåg, Reidun
(2003).
Engineering the stability and oligomerization state of a tetrameric malate dehydrogenase.
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Bjørk, Alexandra; Dalhus, Bjørn; Mantzilas, Dimitrios; Eijsink, Vincent G. H. & Sirevåg, Reidun
(2003).
Malate dehydrogenase from mesophilic and thermophilic green bacteria.
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Benestad, Ingun; Ytteborg, Elisabeth; Vanberg, Christin & Sirevåg, Reidun
(2003).
Two operons for chaperones in Chloroflexus aurantiacus.
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Eijsink, Vincent; Bjørk, Alexandra; Dalhus, Bjørn; Mantzilas, Dimitris & Sirevåg, Reidun
(2002).
Engineering the stability and oligomerization state of a tetrameric malate dehydrogenase.
-
Bjørk, Alexandra; Dalhus, Bjørn; Mantzilas, Dimitris; Eijsink, Vincent & Sirevåg, Reidun
(2002).
Stabilization of tetrameric malate dehydrogenase from Chloroflexus aurantiacus by engineering an ionic network in the dimer-dimer interface.
-
Dalhus, Bjørn; Bjørk, Alexandra; Sirevåg, Reidun & Eklund, Hans
(2002).
Structural basis for protein thermostability - crystal structures of thermostable tetrameric malate dehydrogenases.
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Bjørk, Alexandra; Dalhus, Bjørn; Mantzilas, Dimitrios; Eijsink, Vincent G. H. & Sirevåg, Reidun
(2002).
Engineering the stability and oligomerization state of a tetrameric malate dehydrogenase.
-
Bjørk, Alexandra; Dalhus, Bjørn; Mantzilas, Dimitrios; Eijsink, Vincent G. H. & Sirevåg, Reidun
(2002).
STABILIZATION OF TETRAMERIC MALATE DEHYDROGENASE FROM CHLOROFLEXUS AURANTIACUS BY ENGINEERING AN IONIC NETWORK IN THE DIMER-DIMER INTERFACE.
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Bjørk, Alexandra; Mantzilas, Dimitrios; Dalhus, Bjørn; Eijsink, Vincent G. H. & Sirevåg, Reidun
(2002).
STABILIZATION OF TETRAMERIC MALATE DEHYDROGENASE FROM CHLOROFLEXUS AURANTIACUS BY ENGINEERING AN IONIC NETWORK IN THE DIMER-DIMER INTERFACE.
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Dalhus, Bjørn; Bjørk, Alexandra; Eijsink, Vincent G. H.; Mantzilas, Dimitrios & Sirevåg, Reidun
(2002).
Analysis of an ionic network within a protein oligomer and engeneering towards higher thermostability.
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Bjørk, Alexandra; Dalhus, Bjørn; Mantzilas, Dimitris; Eijsink, Vincent & Sirevåg, Reidun
(2001).
Mutations at the dimer-dimer interface in tetrameric malate dehydrogenase from Chloroflexus aurantiacus have large effects on thermal stability.
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Vanberg, Christin; Bjørk, Alexandra & Sirevåg, Reidun
(2001).
The moderate thermophilic green gliding bacterium Chloroflexus aurantiacus contains to alleles for chaperonins.
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Dalhus, Bjørn; Bjørk, Alexandra; Sirevåg, Reidun & Eklund, Hans
(2001).
Crystal structure of MDHs from thermophilic bacteria - structural basis for thermophilic protein stability.
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Bjørk, Alexandra; Dalhus, Bjørn; Mantzilas, Dimitrios; Eijsink, Vincent G. H. & Sirevåg, Reidun
(2001).
Mutations at the dimer-dimer interface in tetrameric malate dehydrogenase from Chloroflexus aurantiacus have large effects on thermal stability.
-
Dalhus, Bjørn; Bjørk, Alexandra; Eklund, Hans & Sirevåg, Reidun
(2001).
Crystal structures of MDHs from thermophilic bacteria: structural basis for protein thermostability.
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Bjørk, Alexandra; Eijsink, Vincent G. H. & Sirevåg, Reidun
(2000).
Thermostability of tetrameric malate dehydrogenase from the green, gliding bacterium Chloroflexus aurantiacus.
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Vanberg, Christin; Bjørk, Alexandra & Sirevåg, Reidun
(2000).
Molecular chaperones from the moderate thermophilic, green gliding bacterium Chloroflexus aurantiacus.
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Molden, Tor; Bjørk, Alexandra; Tooming, Ave & Sirevåg, Reidun
(1999).
Thermostability in Chloroflexus aurantiacus.
Show summary
In order to investigate the molecular basis of thermostability in bacteria growing at high temperatures, we have isolated and characterized malate dehydrogenase (MDH) and its corresponding gene from several mesophilic and thermophilic phototrophic bacteria. The mdh gene from the moderate thermophile Chloroflexus aurantiacus has been isolated, characterized, cloned and overexpressed in Escherichia coli , and the 3-dimensional structure of the purified and crystallized protein thus produced has been determined. The native enzyme is a tetramer of identical subunits each with a molecular mass of 32.7 kDa. The enzyme is stable at its optimum temperature 55oC, but after 9 min at 65oC, the activity is reduced to half.
An earlier observation in this laboratory which indicated that MDH exists as a tetramer only when the temperature is approaching the optimum temperature, has prompted us to examine the role of specific amino acids in subunit interactions as well as in thermostability. By in vitro mutagenesis, the four mutants W39A, R173N, R173Q and Q188H of mdh from C. aurantiacus have been constructed and expressed in E. coli.. The resulting mutant MDHs have been tested for activity, thermostability and ability to form tetramers. The results obtained will be presented and discussed.
The possibility that cytoplasmic factors like molecular chaperones might be involved in the observed thermostability was examined. In addition to prevent aggregation of newly synthesized non-native polypeptides, chaperones are believed to prevent aggregation of proteins that unfold upon exposure of cells to stress, such as high temperature. When cell free extracts from C. aurantiacus grown at 45oC, 55oC, 60oC and 65oC were analyzed by SDS-PAGE, three distinct bands which increased in intensity with the growth temperature were detected. The bands corresponded to sizes of 11.2 kD, 60 kD and 75 kD and were detected also after heat treatment of the cell free extracts (60 min at 65oC) indicating that they are heat stable. The 60 kD band was eluted from the gel, purified and the sequence of the first 45 amino acids in the N-terminal end determined. Although no cross reaction occurred with antibodies directed against HSP 60 (GroEL) from Escherichia coli, the sequence showed an identity of 55.6% and a similarity of 76% with GroEL. A dendogram which shows the similarity of the present N-terminal sequence to the N-terminal sequences of HSP60 from 12 other organisms will be presented.
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Tooming, Ave & Sirevåg, Reidun
(1999).
Induction of molecular chaperones in the thermophilic green bacterium Chloroflexus aurantiacus.
Show summary
The phototrophic green bacterium C. aurantiacus has an optimum temperature for growth of 55oC, but is capable of growth at temperatures up to 70oC. Earlier observations in our laboratory that addition of cell free extracts from C. aurantiacus increased the thermostability of purified malate dehydrogenase (MDH) from the same organism (1), prompted us to investigate this phenomenon in more detail. It was found that addition of cell free extracts from C. aurantiacus grown at temperatures above 55oC increased the thermostability of purified MDH significantly. Also MDH present in cell free extracts from cells grown at high temperatures was more thermostable than that in extracts from cells grown at low tempoeratures. The possibility that cytoplasmic factors such as molecular chaperones might be involved in maintenance of the observed thermostability was examined. Molecular chaperones are currently defined in purely fuctional terms as proteins that assist in the non-covalent assembly and/or disassembly of protein containig structures in vivo, but ar not componets of these structures when they are performing their normal biological functions (2). Molecular chaperones prevent protein aggregation by binding to exposed hydrophobic residues of newly synthesized non-native polypeptides. In addition, chaperones are believed to prevent aggregation in the case of proteins that unfold upon exposure of cells to stress, such as high temperature.
Cell free extracts of C. aurantiacus grown at 45oC, 55oC, 60oC and 65oC were analysed by SDS-PAGE. Three distinct bands which increased in intensity with growth temperature were detected. These had molecular weights of 11.2 kDa, 60 kDa and 75 kDa and were present after heat treatment of the cell free extracts (60 min at 65oC), indicating that they are thermostable. When the gel was analysed by Western blotting, using antibodies directed against chaperonin hsp60 (GroEL) from Escherichia coli, no reaction occurred. However, when the 60 kDa band was eluted from the gel, purified and the first 45 amino acids of the N-terminal end sequenced, they showed a sequence identity of 55.6% and a similarity of 76% with GroEL from E. coli. A dendrogram will be presented which shows the sequence similarity of the present N-terminal end to the N-terminal ends of cnp60 from 12 other organisms.
(1) Synstad, B., Emmerhoff, O., Sirevåg, R., 1996. Arch Microbiol 165, 346-543.
(2) Ellis, R. J. 1977. Biochem Biophys Res Commun 238, 687-692.
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Bjørk, Alexandra; Molden, Tor; Torgunrud, Annette & Sirevåg, Reidun
(1999).
Thermostable malate dehydrogenases from phototrophic bacteria.
Show summary
In order to investigate the molecular basis of thermostability in proteins, we have isolated and characterized malate dehydrogenase (MDH) and its corresponding gene from mesophilic and thermophilic phototrophic bacteria. The green gliding bacterium Chloroflexus aurantiacus is a moderate thermophile which grows optimally at 55oC. The gene mdh coding for MDH from C. aurantiacus has been isolated characterized, cloned and overexpressed in Escherichia coli (1). The protein thus produced has been purified, crystallized and its 3-dimensional structure determined The native enzyme is a tetramer of identical subunits each with a molecular mass of 32.7 kDa . The enzyme is stable at 55oC, whereas at 65oC, the activity is reduced to the half after 9 min.
An earler observation in this laboratory that tetramers are formed only when the temperature is approaching the optimum temperature of 55oC (2), has prompted us to examine specific amino acids in MDH for their role in subunit interactions as well as in thermostability. By in vitro mutagenesis, the following four mutants of mdh from C. aurantiacus have been constructed and then expressed in E. coli. : W39A, R173N, R173Q and Q188H. In each case, the overproduced protein have been tested for activity and thermostability and the results will be presented and discussed.
Analysis and comparison of corresponding proteins from mesophilic and thermophilic organisms might give valuable information about amino acids important for thermostability and for subunit interactions. For this reason, MDH from the mesophilioc phototrophic bacterium Helibacterium gestii has been purified and characterized. Although H. gestii is a mesophile with an optimum temperature of growth of 37oC, its MDH is remarably thermostable. Thus, at 60oC, the half life, T1/2 of the enzyme is 20 min, whereas T1/2 for MDH from another mesophilic bacterium such as Chlorobium virioforme is 0.5 min at this temperature. The sequence of the 29 first amino acid in the N-terminal end of MDH from H. gestii has been determined and used to construct an oligomer which is used as a probe in order to find the mdh gene in a genomic library made in lambda-ZAP..
(1) Synstad et al. Arch Microbiol 165: 346-353 (1996)
(2) Rolstad et al. J Bacteriol 170: 2947-2953 (1988)
(3) Charnock et al. J Bacteriol 174: 1307-1313 (1992)
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Molden, Tor; Bjørk, Alexandra; Tooming, Ave & Sirevåg, Reidun
(1999).
Thermostability in Chloroflexus aurantiacus.
Show summary
In order to investigate the molecular basis of thermostability in bacteria growing at high temperatures, we have isolated and characterized malate dehydrogenase (MDH) and its corresponding gene from several mesophilic and thermophilic phototrophic bacteria. The mdh gene from the moderate thermophile Chloroflexus aurantiacus has been isolated, characterized, cloned and overexpressed in Escherichia coli , and the 3-dimensional structure of the purified and crystallized protein thus produced has been determined. The native enzyme is a tetramer of identical subunits each with a molecular mass of 32.7 kDa. The enzyme is stable at its optimum temperature 55oC, but after 9 min at 65oC, the activity is reduced to half.
An earlier observation in this laboratory which indicated that MDH exists as a tetramer only when the temperature is approaching the optimum temperature, has prompted us to examine the role of specific amino acids in subunit interactions as well as in thermostability. By in vitro mutagenesis, the four mutants W39A, R173N, R173Q and Q188H of mdh from C. aurantiacus have been constructed and expressed in E. coli.. The resulting mutant MDHs have been tested for activity, thermostability and ability to form tetramers. The results obtained will be presented and discussed.
The possibility that cytoplasmic factors like molecular chaperones might be involved in the observed thermostability was examined. In addition to prevent aggregation of newly synthesized non-native polypeptides, chaperones are believed to prevent aggregation of proteins that unfold upon exposure of cells to stress, such as high temperature. When cell free extracts from C. aurantiacus grown at 45oC, 55oC, 60oC and 65oC were analyzed by SDS-PAGE, three distinct bands which increased in intensity with the growth temperature were detected. The bands corresponded to sizes of 11.2 kD, 60 kD and 75 kD and were detected also after heat treatment of the cell free extracts (60 min at 65oC) indicating that they are heat stable. The 60 kD band was eluted from the gel, purified and the sequence of the first 45 amino acids in the N-terminal end determined. Although no cross reaction occurred with antibodies directed against HSP 60 (GroEL) from Escherichia coli, the sequence showed an identity of 55.6% and a similarity of 76% with GroEL. A dendogram which shows the similarity of the present N-terminal sequence to the N-terminal sequences of HSP60 from 12 other organisms will be presented.
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Karlsen, Frank; Andreassen, Trine-Merethe; Rabone, Marian; Skrodenyte, Vesta; Sirevåg, Reidun & Eckner, Karl
[Show all 10 contributors for this article]
(1999).
Development of molecular methods for rapid detection of coliform bacteria in water.
Show summary
The microbiological examination of drinking water has traditionally been performed by techniques such as membrane filtration (MF) and most probable number (MPN), using coliform bacteria as an indicator of fecal contamination. However, new methods such as those based on defined substrate technology are beginning to be accepted as well. Based on fluorescence measurements to assay the activity of b-galactosidase, Colifast ASA has developed an istrument, CA100 which automatically detects E. coli after 7-9 hours of growth in a specific enrichment medium. In addition, molecular methods based on the increasing knowledge of nucleic acid sequences, as well as immunological techniques, are being developed in several laboratories.
In the present project, the enzymatic method developed by Colifast was compared with three molecular methods developed in our laboratory at UiO. For comparison and control purpose, traditional plate counts, biochemical tests and API 20E (Biomerieux) were included in the experiments. The first and second method utilise PCR (Polymerase Chain Reaction) after concentration of the samples by either centrifugation or by the aid of immunomagnetic beads coated with antibodies directed against E. coli. For identification of E. coli, a primer-pair was constructed which was shown to be E. coli specific both by homology search using Blast 2.0 (NCBI, USA) and by testing with PCR against 20 other commmon bacterial species. As a control of the the DNAS-quality, a universal primer-pair directed against a conserved region of the 16S rRNA gene was constructed and used in each experiment. The third method is a sandwich-ELISA (Enzyme-linked Immunoasorbent Assay) test, using antibodies against b-galactosidase. Samples included water from the Lysaker river as well as reconstituted freeze-dried reference cultures of defined composition.
