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Literature summary for 3.1.26.11 extracted from

  • Spaeth, B.; Settele, F.; Schilling, O.; DAngelo, I.; Vogel, A.; Feldmann, I.; Meyer-Klaucke, W.; Marchfelder, A.
    Metal requirements and phosphodiesterase activity of tRNase Z enzymes (2007), Biochemistry, 46, 14742-14750.
    View publication on PubMed

Application

Application Comment Organism
analysis comparative studies both bis(p-nitrophenyl)phosphate hydrolysis and pre-tRNA processing of tRNase Z variants Arabidopsis thaliana

Cloned(Commentary)

Cloned (Comment) Organism
expressed in Escherichia coli Drosophila melanogaster
expressed in Escherichia coli Saccharomyces cerevisiae
expressed in Escherichia coli, prior to electrophoretic mobility shift assay (EMSA) and cross-linking experiments Arabidopsis thaliana

Protein Variants

Protein Variants Comment Organism
C25G variants of AthTRZ1 wild-type enzyme generated by mutagenesis Arabidopsis thaliana
C40G variants of AthTRZ1 wild-type enzyme generated by mutagenesis Arabidopsis thaliana
E208A variants of AthTRZ1 wild-type enzyme generated by mutagenesis Arabidopsis thaliana
F51L variants of AthTRZ1 wild-type enzyme generated by mutagenesis Arabidopsis thaliana
G62V variants of AthTRZ1 wild-type enzyme generated by mutagenesis Arabidopsis thaliana
P178A variants of AthTRZ1 wild-type enzyme generated by mutagenesis Arabidopsis thaliana
P64A variants of AthTRZ1 wild-type enzyme generated by mutagenesis Arabidopsis thaliana
R252G variants of AthTRZ1 wild-type enzyme generated by mutagenesis Arabidopsis thaliana

Inhibitors

Inhibitors Comment Organism Structure
bis(p-nitrophenyl)phosphate pre-tRNA processing inhibited by bis(p-nitrophenyl)phosphate concentrations higher than 10 mM observed Drosophila melanogaster
bis(p-nitrophenyl)phosphate pre-tRNA processing inhibited by bis(p-nitrophenyl)phosphate concentrations higher than 10 mM observed Saccharomyces cerevisiae
additional information reaction performed with 20 microM bis(p-nitrophenyl)phosphate and 20 microM tRNA, bis(p-nitrophenyl)phosphate does not inhibit tRNA binding Arabidopsis thaliana

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
1.2
-
bis(p-nitrophenyl)phosphate AthTRZ1-P178A, mutant, bis(p-nitrophenyl)phosphate catalysis Arabidopsis thaliana
1.8
-
bis(p-nitrophenyl)phosphate AthTRZ1-R252G, mutant, bis(p-nitrophenyl)phosphate catalysis Arabidopsis thaliana
3.5
-
bis(p-nitrophenyl)phosphate AthTRZ1-E208A, mutant, bis(p-nitrophenyl)phosphate catalysis Arabidopsis thaliana
6
-
bis(p-nitrophenyl)phosphate AthTRZ1-C40G, mutant, bis(p-nitrophenyl)phosphate catalysis Arabidopsis thaliana
6.5
-
bis(p-nitrophenyl)phosphate AthTRZ1-F51L, mutant, bis(p-nitrophenyl)phosphate catalysis Arabidopsis thaliana
8
-
bis(p-nitrophenyl)phosphate AthTRZ1-G62V, mutant, bis(p-nitrophenyl)phosphate catalysis Arabidopsis thaliana
8.5
-
bis(p-nitrophenyl)phosphate AthTRZ1 wild-type, hydrolysis of phosphodiester bonds of bpNPP, KM value for AthTRZ1 two-fold increased relative to that of the tRNase Z of Escherichia coli Arabidopsis thaliana
13.2
-
bis(p-nitrophenyl)phosphate AthTRZ1-C25G, mutant, bis(p-nitrophenyl)phosphate catalysis Arabidopsis thaliana
22.2
-
bis(p-nitrophenyl)phosphate AthTRZ1-P64A, mutant, bis(p-nitrophenyl)phosphate catalysis Arabidopsis thaliana

Metals/Ions

Metals/Ions Comment Organism Structure
Ca2+ requird for in Vitro tRNA processing reaction Arabidopsis thaliana
Mg2+ 2 mM, also required for in vitro tRNA processing reactions Arabidopsis thaliana
Mn2+ 0.2 mM, addition of Mn2+ ions triples activity, also required for in Vitro tRNA processing reactions Arabidopsis thaliana
additional information in contrast to bis(p-nitrophenyl)phosphate hydrolysis, pre-tRNA processing requires additional metal ions, Mn2+ or Mg2+, as Zn2+ ions alone are insufficient, metal dependence of the in vitro processing reaction analyzed, bis(p-nitrophenyl)phosphate activity of the chelator-treated AthTRZ1 without (TRZ-E) and with additional metal ions tested, chelator-treated AthTRZ1 without additional metal ions reveal same activity as the nonchelator-treated AthTRZ1 Arabidopsis thaliana
Zn2+ 0.2 mM, addition of Zn2+ ions to the chelator-treated enzyme doubles activity, high affinity to Zn2+ even upon incubation with metal chelators, 0.76 Zn2+ ions retained per dimer Arabidopsis thaliana

Organism

Organism UniProt Comment Textmining
Arabidopsis thaliana
-
AthTRZ1, wild-type and mutant variants studied
-
Drosophila melanogaster
-
-
-
Saccharomyces cerevisiae P36159 long tRNase Z enzyme
-

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
additional information
-
in vitro tRNA processing assays, bis(p-nitrophenyl)phosphate hydrolysis assay described, in contrast to the short tRNase Z enzyme AthTRZ1 of Arabidopsis thaliana, long tRNase Z enzymes do not have bis(p-nitrophenyl)phosphate hydrolysis activity Saccharomyces cerevisiae
additional information
-
in vitro tRNA processing assays, bis(p-nitrophenyl)phosphate hydrolysis assay described, kinetic analysis shown, AthTRZ1 effective catalyst of bpNPP hydrolysis, AthTRZ1-R252G reveals ten times higher activity compared to the wild-type enzyme, in contrast to the short tRNase Z enzyme AthTRZ1, long tRNase Z enzymes do not have bis(p-nitrophenyl)phosphate hydrolysis activity Arabidopsis thaliana
additional information
-
in vitro tRNA processing assays, bpNPP hydrolysis assay described, in contrast to the short tRNase Z enzyme AthTRZ1 of Arabidopsis thaliana, long tRNase Z enzymes do not have bis(p-nitrophenyl)phosphate hydrolysis activity Drosophila melanogaster

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
bis(p-nitrophenyl)phosphate + H2O differentiation of tRNase Z variants by 3'-processing activity and their ability to hydrolyze the phosphodiester bond in the chromogenic phosphodiester bis(p-nitrophenyl)phosphate (bpNPP), smallest known tRNase Z substrate, fourteen variants lost ability to hydrolyze bpNPP, seven variants reveal reduced activity Arabidopsis thaliana p-nitrophenol + p-nitrophenyl phosphate
-
?
bis(p-nitrophenyl)phosphate + H2O no hydrolysis of the the chromogenic phosphodiester bis(p-nitrophenyl)phosphate (bpNPP) observed Drosophila melanogaster p-nitrophenol + p-nitrophenyl phosphate
-
?
bis(p-nitrophenyl)phosphate + H2O no hydrolysis of the the chromogenic phosphodiester bis(p-nitrophenyl)phosphate (bpNPP) observed Saccharomyces cerevisiae p-nitrophenol + p-nitrophenyl phosphate
-
?

Subunits

Subunits Comment Organism
dimer
-
Arabidopsis thaliana

Synonyms

Synonyms Comment Organism
AthTrz1
-
Arabidopsis thaliana
tRNase Z
-
Drosophila melanogaster
tRNase Z
-
Arabidopsis thaliana
tRNase Z
-
Saccharomyces cerevisiae

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
additional information
-
additional information turnover rate of AthTRZ1 eight-fold reduced if compared to tRNAse Z of Escherichia coli Arabidopsis thaliana