PPR65 - Plant Editosome Database - BIG Data Center

Summary

Editing Factor: PPR65
Synonym: NA
Description: Purified, recombinant PPR65 exhibits robust editase activity on synthetic RNAs containing cognate, mitochondrial PpccmFC sequences in vitro, indicating that a PPR protein with a DYW domain is solely sufficient for catalyzing C-to-U RNA editing in vitro; Inductively coupled plasma (ICP)-MS analysis indicated a stoichiometry of two zinc ions per highly active PPR65 monomer; Editing activity was sensitive to addition of zinc acetate or the zinc chelators 1,10-o-phenanthroline and EDTA. Addition of ATP or nonhydrolyzable nucleotide analogs stimulated PPR65-catalyzed RNA-editing activity on PpccmFC substrates, indicating potential allosteric regulation of PPR65 by ATP; RNA oligonucleotides with a single incorporated zebularine also did not disrupt editing in vitro, suggesting that PPR65 cannot bind modified bases due to differences in the structure of the active site compared with other zinc-dependent nucleotide deaminases
Protein Family: PPR
Subclass: DYW
Construct Structure: NA
Gene ID & Species: NA (Physcomitrium patens)
Edited Gene(s): ccmFc
Editing Type(s): C-to-U (45)
Publication(s): [1] A plant pentatricopeptide repeat protein with a DYW-deaminase domain is sufficient for catalyzing C-to-U RNA editing ., The Journal of biological chemistry, 2020. [PMID=31996373]

Editing Details

Species Gene ID Organelle Edited Gene Position Region Editing Type Codon Amino Acid Molecular Effect Experiment Details
Physcomitrium patens NA Mitochondrion ccmFc NA NA C-to-U NA=>NA NA=>NA NA
Experiment Details
Genotype (Ecotype) Allele Treatment Treatment Detail Mutant Type Phenotype Tissue Development Stage Detection Method Editing Frequency Editing Extent Mutant Effect PMID
NAWTEditing reactions assayed with varying ATP concentrationsWith 0 mM ATPNon-mutantNANANAPoisoned primer extension (PPE) assay23.33%LowNone31996373
NAWTEditing reactions assayed with varying ATP concentrationsWith 1 mM ATPNon-mutantNANANAPoisoned primer extension (PPE) assay30.00%LowNone31996373
NAWTEditing reactions assayed with varying ATP concentrationsWith 2 mM ATPNon-mutantNANANAPoisoned primer extension (PPE) assay28.00%LowNone31996373
NAWTEditing reactions assayed with varying ATP concentrationsWith 5 mM ATPNon-mutantNANANAPoisoned primer extension (PPE) assay26.88%LowNone31996373
NAWTEditing reactions assayed with varying Mg++ concentrationsWith 0 mM Mg++Non-mutantNANANAPoisoned primer extension (PPE) assay25.00%LowNone31996373
NAWTEditing reactions assayed with varying Mg++ concentrationsWith 1.5 mM Mg++Non-mutantNANANAPoisoned primer extension (PPE) assay34.00%LowNone31996373
NAWTEditing reactions assayed with varying Mg++ concentrationsWith 3 mM Mg++Non-mutantNANANAPoisoned primer extension (PPE) assay35.00%LowNone31996373
NAWTEditing reactions assayed with varying Mg++ concentrationsWith 6 mM Mg++Non-mutantNANANAPoisoned primer extension (PPE) assay27.00%LowNone31996373
NAWTEditing reactions assayed with varying temperature conditions10 ℃Non-mutantNANANAPoisoned primer extension (PPE) assay0.50%PoorNone31996373
NAWTEditing reactions assayed with varying temperature conditions16 ℃Non-mutantNANANAPoisoned primer extension (PPE) assay1.00%PoorNone31996373
NAWTEditing reactions assayed with varying temperature conditions22 ℃Non-mutantNANANAPoisoned primer extension (PPE) assay12.00%PoorNone31996373
NAWTEditing reactions assayed with varying temperature conditions28 ℃Non-mutantNANANAPoisoned primer extension (PPE) assay24.00%LowNone31996373
NAWTEditing reactions assayed with varying temperature conditions34 ℃Non-mutantNANANAPoisoned primer extension (PPE) assay10.00%PoorNone31996373
NAWTEditing reactions assayed with varying temperature conditions40 ℃Non-mutantNANANAPoisoned primer extension (PPE) assay0.50%PoorNone31996373
NAWTProtein peaks from rPPR65 purificationPeak ANon-mutantNANANAPoisoned primer extension (PPE) assay35.00%LowNone31996373
NAWTProtein peaks from rPPR66 purificationPeak BNon-mutantNANANAPoisoned primer extension (PPE) assay60.00%HighNone31996373
NAWTProtein peaks from rPPR67 purificationPeak CNon-mutantNANANAPoisoned primer extension (PPE) assay18.00%PoorNone31996373
NAWTAddition of zinc chelatorControlNon-mutantNANANAPoisoned primer extension (PPE) assay65.00%HighNone31996373
NAWTAddition of zinc chelatorAddition of 8 mM 1,10-o-phenantroline (PHEN)Non-mutantNANANAPoisoned primer extension (PPE) assay30.00%LowNone31996373
NAWTAddition of zinc chelatorAddition of 8 mM EDTANon-mutantNANANAPoisoned primer extension (PPE) assay36.00%LowNone31996373
NAWTAddition of ATP and non-hydrolyzable analog0 mM ATPNon-mutantNANANAPoisoned primer extension (PPE) assay53.00%MediumNone31996373
NAWTAddition of ATP and non-hydrolyzable analog2mM ATPNon-mutantNANANAPoisoned primer extension (PPE) assay64.00%HighNone31996373
NAWTAddition of ATP and non-hydrolyzable analog2mM ADPNon-mutantNANANAPoisoned primer extension (PPE) assay54.00%MediumNone31996373
NAWTAddition of ATP and non-hydrolyzable analog2mM AMPNon-mutantNANANAPoisoned primer extension (PPE) assay51.00%MediumNone31996373
NAWTAddition of ATP and non-hydrolyzable analog2mM AMP-PCPNon-mutantNANANAPoisoned primer extension (PPE) assay61.00%HighNone31996373
NAWTAddition of ATP and non-hydrolyzable analog2mM AMP-CPNon-mutantNANANAPoisoned primer extension (PPE) assay64.00%HighNone31996373
NAWTAddition of putative transition state inhibitorControlNon-mutantNANANAPoisoned primer extension (PPE) assay23.00%LowNone31996373
NAWTAddition of putative transition state inhibitorzebularine (ZEB)Non-mutantNANANAPoisoned primer extension (PPE) assay24.00%LowNone31996373
NAWTAddition of putative transition state inhibitordeazacytidine (AZA)Non-mutantNANANAPoisoned primer extension (PPE) assay27.00%LowNone31996373
NAWTAddition of putative transition state inhibitortetrahydrouridine (THU)Non-mutantNANANAPoisoned primer extension (PPE) assay26.00%LowNone31996373
NAWTRNA oligonucleotides with incorporated zebularine (Z) with the sequence shown were added to PPR65 reactions at 1000:1 molar ratio to substrates.ControlNon-mutantNANANAPoisoned primer extension (PPE) assay59.00%MediumNone31996373
NAWTRNA oligonucleotides with incorporated zebularine (Z) with the sequence shown were added to PPR65 reactions at 1000:1 molar ratio to substrates.3-mer UZANon-mutantNANANAPoisoned primer extension (PPE) assay58.00%MediumNone31996373
NAWTRNA oligonucleotides with incorporated zebularine (Z) with the sequence shown were added to PPR65 reactions at 1000:1 molar ratio to substrates.11-mer AUCCUZAAAAANon-mutantNANANAPoisoned primer extension (PPE) assay62.00%HighNone31996373
NAWTCatalyzed reactions at a range of RNA concentrations0.08 nMNon-mutantNANANAPoisoned primer extension (PPE) assay39.00%LowNone31996373
NAWTCatalyzed reactions at a range of RNA concentrations0.16 nMNon-mutantNANANAPoisoned primer extension (PPE) assay40.00%MediumNone31996373
NAWTCatalyzed reactions at a range of RNA concentrations0.40 nMNon-mutantNANANAPoisoned primer extension (PPE) assay37.00%LowNone31996373
NAWTCatalyzed reactions at a range of RNA concentrations0.80 nMNon-mutantNANANAPoisoned primer extension (PPE) assay38.00%LowNone31996373
NAWTCatalyzed reactions at a range of RNA concentrations1.6 nMNon-mutantNANANAPoisoned primer extension (PPE) assay36.00%LowNone31996373
NAWTCatalyzed reactions at a range of RNA concentrations4.0 nMNon-mutantNANANAPoisoned primer extension (PPE) assay35.00%LowNone31996373
NAWTEditing assays performed in vitro using different ratios of enzyme versus substrate (E/S)375:01:00Non-mutantNANANAPoisoned primer extension (PPE) assay70.00%HighNone31996373
NAWTEditing assays performed in vitro using different ratios of enzyme versus substrate (E/S)100:01:00Non-mutantNANANAPoisoned primer extension (PPE) assay53.00%MediumNone31996373
NAWTEditing assays performed in vitro using different ratios of enzyme versus substrate (E/S)10:01Non-mutantNANANAPoisoned primer extension (PPE) assay5.00%PoorNone31996373
NAWTEditing assays performed in vitro using different ratios of enzyme versus substrate (E/S)1:01Non-mutantNANANAPoisoned primer extension (PPE) assayNANANone31996373
NAWTEditing assays performed in vitro using different ratios of enzyme versus substrate (E/S)1:10Non-mutantNANANAPoisoned primer extension (PPE) assayNANANone31996373
NAWTEditing assays performed in vitro using different ratios of enzyme versus substrate (E/S)0.111111111Non-mutantNANANAPoisoned primer extension (PPE) assayNANANone31996373
Last update: Jul 2021 (version 1.0)