Literature summary extracted from
Hardwick, S.W.; Gubbey, T.; Hug, I.; Jenal, U.; Luisi, B.F.
Crystal structure of Caulobacter crescentus polynucleotide phosphorylase reveals a mechanism of RNA substrate channelling and RNA degradosome assembly (2012), Open biology, 2, 120028.
Cloned(Commentary)
EC Number |
Cloned (Comment) |
Organism |
---|
2.7.7.8 |
gene pnp, cloned in a plasmid with rne gene, encoding endoribonuclease RNase E, fragments, expression of N-terminally GST- or His6-tagged enzyme fragments in Escherichia coli |
Caulobacter vibrioides |
Crystallization (Commentary)
EC Number |
Crystallization (Comment) |
Organism |
---|
2.7.7.8 |
purified recombinant RNA-free and RNA-bound PNPase, as protein-peptide and protein-RNA complexes, mixing of PNPase recognition peptide from RNase E, KPRRGWWRR, with apo-PNPase in a 2:1 ratio, sitting drop vapour diffusion method, mixing of equal volumes of protein solution and cyrstallization solution, containing w/v PEG 3350, 0.1 M Bis-Tris, pH 5.5, 0.1 M ammonium acetate for RNA-bound crystals, and with the apo-enzyme containing w/v PEG3000, 0.1 M trisodium citrate, pH 5.5, for hexagonal crystals and w/v PEG 3350, 0.15 M DL-malic acid for rhombohedral crystals, 18°C, 1 week, X-ray diffraction structure determination aand analysis at 2.6-3.3 A resolution, molecular replacement |
Caulobacter vibrioides |
Organism
EC Number |
Organism |
UniProt |
Comment |
Textmining |
---|
2.7.7.8 |
Caulobacter vibrioides |
- |
gene pnp |
- |
2.7.7.8 |
Caulobacter vibrioides NA1000 |
- |
gene pnp |
- |
Purification (Commentary)
EC Number |
Purification (Comment) |
Organism |
---|
2.7.7.8 |
recombinant N-terminally GST-tagged enzyme fragments from Escherichia coli by glutathione affinity chromatography, of His6-tagged enzyme fragments by nickel affinity chromatography, followed by gel filtration in both procedures |
Caulobacter vibrioides |
Subunits
EC Number |
Subunits |
Comment |
Organism |
---|
2.7.7.8 |
trimer |
the enzyme has a ring-like, trimeric architecture that creates a central channel where phosphorolytic active sites reside. One face of the ring is decorated with RNA-binding K-homology (KH) and S1 domains, domain organization with modular organization of conserved structural domains, overview |
Caulobacter vibrioides |
Synonyms
EC Number |
Synonyms |
Comment |
Organism |
---|
2.7.7.8 |
PNPase |
- |
Caulobacter vibrioides |
2.7.7.8 |
polynucleotide phosphorylase |
- |
Caulobacter vibrioides |
Temperature Optimum [°C]
EC Number |
Temperature Optimum [°C] |
Temperature Optimum Maximum [°C] |
Comment |
Organism |
---|
2.7.7.8 |
37 |
- |
assay at |
Caulobacter vibrioides |
pH Optimum
EC Number |
pH Optimum Minimum |
pH Optimum Maximum |
Comment |
Organism |
---|
2.7.7.8 |
7.5 |
- |
assay at |
Caulobacter vibrioides |
General Information
EC Number |
General Information |
Comment |
Organism |
---|
2.7.7.8 |
evolution |
two domains, both resembling closely the phosphorolytic exoribonuclease RNase PH, EC 27.7.56, almost certainly have originated from duplication and fusion of an ancestral gene. While the C-terminal RNase PH-like domain catalyses phosphorolytic attack of RNA, the N-terminal domain has lost this capacity. Instead, it contributes to the ring-like quaternary structure of the trimeric PNPase assembly |
Caulobacter vibrioides |
2.7.7.8 |
additional information |
the enzyme has a ring-like, trimeric architecture that creates a central channel where phosphorolytic active sites reside, with asymmetry within the catalytic core of the enzyme. One face of the ring is decorated with RNA-binding K-homology (KH) and S1 domains. In the RNA-free form, the S1 domains adopt a splayed conformation that may facilitate capture of RNA substrates. In the RNA-bound structure, the three KH domains collectively close upon the RNA and direct the 3' end towards a constricted aperture at the entrance of the central channel. Structural non-equivalence, induced upon RNA binding, helps to channel substrate to the active sites through mechanical ratcheting. Access to the PNPase active sites is through the central channel, which can accommodate single-stranded RNA with some structural adjustment of a constricted aperture at the channel entrance, residues and motifs involved in RNA directionality, recognition, and quarternary changes in the core, structure-function-relationship, detailed overview |
Caulobacter vibrioides |
2.7.7.8 |
physiological function |
polynucleotide phosphorylase is an exoribonuclease that cleaves single-stranded RNA substrates with 3' -5' directionality and processive behaviour |
Caulobacter vibrioides |