The only exoribonuclease present in Haloferax volcanii has an unique response to temperature changes.
BACKGROUND
Little is known regarding mRNA degradation mechanisms in archaea. In some of these single-cell organisms the existence of a complex of exoribonucleases called the exosome has been demonstrated. However, in halophilic archaea the RNase R homologue is essential since it is the only enzyme described with exoribonucleolytic activity.


METHODS
In this work we have characterized the mechanism of action of Haloferax volcanii RNase R and its implications for the RNA degradation process. We have determined the salt, pH and divalent ion preference, and set the best conditions for the activity assays. Furthermore, we have determined the activity of the protein at different temperatures using different substrates. The dissociation constants were also calculated by Surface Plasmon Resonance. Finally, we have built a model and compared it with the Escherichia coli counterparts.


RESULTS
The results obtained showed that at 37°C, in spite of being named RNase R, this protein behaves like an RNase II protein, halting when it reaches secondary structures, and releasing a 4 nt end-product. However, at 42°C, the optimum temperature of growth, this protein is able to degrade secondary structures, acting like RNase R.


CONCLUSIONS
This discovery has a great impact for RNA degradation, since this is the first case reported where a single enzyme has two different exoribonucleolytic activities according to the temperature. Furthermore, the results obtained are very important to help to decipher the RNA degradation mechanisms in H. volcanii, since RNase R is the only exoribonuclease involved in this process.
Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa, Av da República, 2780-157 Oeiras, Portugal.
Rute G
RG
Matos
Eduardo
E
López-Viñas
Paulino
P
Goméz-Puertas
Cecília M
CM
Arraiano
D000595:Amino Acid Sequence; D002384:Catalysis; D004789:Enzyme Activation; D005095:Exoribonucleases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D019622:Haloferax volcanii / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D020871:RNA Stability / Q000502:physiology; D012330:RNA, Double-Stranded / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013696:Temperature
D012330:RNA, Double-Stranded; D005095:Exoribonucleases; C080868:ribonuclease R