PMID:15705581

Citation	Purusharth, RI, Klein, F, Sulthana, S, Jäger, S, Jagannadham, MV, Evguenieva-Hackenberg, E, Ray, MK and Klug, G (2005) Exoribonuclease R interacts with endoribonuclease E and an RNA helicase in the psychrotrophic bacterium Pseudomonas syringae Lz4W. J. Biol. Chem. 280:14572-8
Abstract	Endoribonuclease E, a key enzyme involved in RNA decay and processing in bacteria, organizes a protein complex called degradosome. In Escherichia coli, Rhodobacter capsulatus, and Streptomyces coelicolor, RNase E interacts with the phosphate-dependent exoribonuclease polynucleotide phosphorylase, DEAD-box helicase(s), and additional factors in an RNA-degrading complex. To characterize the degradosome of the psychrotrophic bacterium Pseudomonas syringae Lz4W, RNase E was enriched by cation exchange chromatography and fractionation in a glycerol density gradient. Most surprisingly, the hydrolytic exoribonuclease RNase R was found to co-purify with RNase E. Co-immunoprecipitation and Ni(2+)-affinity pull-down experiments confirmed the specific interaction between RNase R and RNase E. Additionally, the DEAD-box helicase RhlE was identified as part of this protein complex. Fractions comprising the three proteins showed RNase E and RNase R activity and efficiently degraded a synthetic stem-loop containing RNA in the presence of ATP. The unexpected association of RNase R with RNase E and RhlE in an RNA-degrading complex indicates that the cold-adapted P. syringae has a degradosome of novel structure. The identification of RNase R instead of polynucleotide phosphorylase in this complex underlines the importance of the interaction between endo- and exoribonucleases for the bacterial RNA metabolism. The physical association of RNase E with an exoribonuclease and an RNA helicase apparently is a common theme in the composition of bacterial RNA-degrading complexes.
Links	PubMed Online version:10.1074/jbc.M413507200
Keywords	Adenosine Triphosphate/chemistry; Blotting, Western; Centrifugation, Density Gradient; Chromatography, Ion Exchange; Electrophoresis, Polyacrylamide Gel; Endoribonucleases/chemistry; Endoribonucleases/metabolism; Endoribonucleases/physiology; Exoribonucleases/metabolism; Glycerol/pharmacology; Immunoprecipitation; Protein Binding; Pseudomonas syringae/enzymology; RNA/chemistry; RNA/metabolism; RNA Helicases/metabolism; Recombinant Proteins/chemistry; Ribonucleases/chemistry; Time Factors; Transcription, Genetic
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