3.1.26.5: ribonuclease P
This is an abbreviated version!
For detailed information about ribonuclease P, go to the full flat file.
Word Map on EC 3.1.26.5
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3.1.26.5
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ribonucleoproteins
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pre-trnas
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endoribonuclease
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endonucleolytic
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eculizumab
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aminoacylation
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rna-based
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stem-loops
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phosphodiester
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nucleolar
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nocturnal
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eubacterial
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paroxysmal
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gene-targeting
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anticodon
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base-pairing
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rna-protein
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sars-cov-2
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c5a
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horikoshii
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hemoglobinuria
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trna-like
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pyrococcus
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swab
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3'-terminal
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5\'-leader
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riboswitches
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trnatyr
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2'-hydroxyls
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trnahis
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trailer
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cloverleaf
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rna-rna
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self-splicing
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protein-rna
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tmrna
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micrococcal
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self-cleaving
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t-loops
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p-mediated
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complement-mediated
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ncrnas
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pseudoknots
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watson-crick
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eukaryal
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oligoribonucleotides
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analysis
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pharmacology
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snornps
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synthesis
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medicine
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biotechnology
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trnase
- 3.1.26.5
- ribonucleoproteins
- pre-trnas
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endoribonuclease
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endonucleolytic
- eculizumab
- aminoacylation
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rna-based
-
stem-loops
-
phosphodiester
-
nucleolar
-
nocturnal
-
eubacterial
-
paroxysmal
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gene-targeting
-
anticodon
-
base-pairing
-
rna-protein
- sars-cov-2
- c5a
- horikoshii
- hemoglobinuria
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trna-like
- pyrococcus
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swab
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3'-terminal
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5\'-leader
- riboswitches
- trnatyr
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2'-hydroxyls
- trnahis
- trailer
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cloverleaf
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rna-rna
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self-splicing
-
protein-rna
- tmrna
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micrococcal
-
self-cleaving
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t-loops
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p-mediated
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complement-mediated
- ncrnas
-
pseudoknots
-
watson-crick
-
eukaryal
- oligoribonucleotides
- analysis
- pharmacology
-
snornps
- synthesis
- medicine
- biotechnology
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trnase
Reaction
endonucleolytic cleavage of RNA, removing 5'-extranucleotides from tRNA precursor =
Synonyms
Aq_880, aRpp29, aRpp29 protein, AtPop1p, AtPRORP1, AtPRORP2, AtPRORP3, C5 protein, CrPRORP, hPOP1, hPOP4, hPOP7, M1 RNA, M1GS, M1GS RNA, mitochondrial RNase P protein 1, MRPP1, MRPP2, MRPP3, nuclear ribonclease P ribonucleoprotein, nuclease, ribo-, P, Pfu Pop5, PhoPRNA, POP1, Pop1p, Pop5, Pop6, Pop7, PRORP, PRORP1, PRORP2, PRORP3, Protein C5, protein-only ribonuclease P, protein-only RNase P, protein-only RNase P enzyme, proteinaceous RNase P, ribonuclease MRP, ribonuclease P, ribonuclease P ribozyme, ribosomal RNA processing ribonucleoprotein, Ribunuclease P, RNA processing protein POP1, RNA processing protein POP5, RNA processing protein POP6, RNA processing protein POP7, RNA processing protein POP8, RNase MRP, RNase P, RNase P holoenzyme, RNase P protein, RNase P ribozyme, RNase P RNA, RNase P/MRP, RNase P/MRP protein, RNaseP protein, RNaseP protein p20, RNaseP protein p30, RNaseP protein p38, RNaseP protein p40, RNases P, RNP, Rpm2p, RPP, RPP14, Rpp20, Rpp21, Rpp25, Rpp29, Rpp30, Rpp38, RPP40, RPR, RPR1, transfer RNA 5' maturation enzyme, transfer RNA processing enzyme, tRNA processing enzyme, tRNA-processing endonuclease, tRNA-processing enzyme
ECTree
3.1.26.5 ribonuclease PAdvanced search results
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results (Engineering
Engineering on EC 3.1.26.5 - ribonuclease P
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PROTEIN VARIANTS 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
C344A
site-directed mutagenesis, the mutant shows a 19% reduced zinc level compared to the wild-type enzyme
C347A
site-directed mutagenesis, the mutant shows a 29% reduced zinc level compared to the wild-type enzyme
C565A
site-directed mutagenesis, the mutant shows a 75% reduced zinc level compared to the wild-type enzyme
C56A
site-directed mutagenesis, the mutant shows no enzyme activity
C56G
site-directed mutagenesis, the mutant shows no enzyme activity
C57A
site-directed mutagenesis, the mutant shows 90% of wild-type enzyme activity
C57C
site-directed mutagenesis, the mutant shows 10% of wild-type enzyme activity
D399A
mutation significantly reduces activity. No product formation is observed after a 30-min incubation under standard STO assay conditions (1 mM MgCl2). No increase in activity, when Mg2+ concentration increases from 1 mM to 20 mM
D474A
mutation significantly reduces activity. No product formation is observed after a 30-min incubation under standard STO assay conditions (1 mM MgCl2). Activity increases significantly at the 20 mM Mg2+ compared to 1 mM Mg2+
D474A/D475A
site-directed mutagenesis, the mutant shows 3,3% reduction of the zinc level compared to the wild-type enzyme
D475A
mutation significantly reduces activity. No product formation is observed after a 30-min incubation under standard STO assay conditions (1 mM MgCl2). Activity increases significantly at the 20 mM Mg2+ compared to 1 mM Mg2+
D493A
mutation significantly reduces activity. No product formation is observed after a 30-min incubation under standard STO assay conditions (1 mM MgCl2). No increase in activity, when Mg2+ concentration increases from 1 mM to 20 mM
G18A
site-directed mutagenesis, the mutant shows 15% of wild-type enzyme activity
G18C
site-directed mutagenesis, the mutant shows 10% of wild-type enzyme activity
G19A
site-directed mutagenesis, the mutant shows 85% of wild-type enzyme activity
G19C
site-directed mutagenesis, the mutant shows 90% of wild-type enzyme activity
H548A
site-directed mutagenesis, the mutant shows a 60% reduced zinc level compared to the wild-type enzyme
E40C
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site-directed mutagenesis, comparison of metal effects on enzyme-substrate complex formation with the wild-type enzyme
F16A
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affinity of RNase P for A(-4) pre-tRNA is decreased more than 10fold
F20A
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decrease in the affinity of A(-4) pre-tRNA compared to that of G(-4) pre-tRNA, causing the A(-4)/G(-4) selectivity ratio to decrease to less than 1. Mutations in the central cleft of P protein alters the observed cleavage rate constant by less than 2fold
K64C
R60A
R62A
R65A
R65C
R68A
R68C
Y113C
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site-directed mutagenesis, comparison of metal effects on enzyme-substrate complex formation with the wild-type enzyme
Y34A
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decrease in the affinity of A(-4) pre-tRNA compared to that of G(-4) pre-tRNA, causing the A(-4)/G(-4) selectivity ratio to decrease to less than 1. Mutations in the central cleft of P protein alters the observed cleavage rate constant by less than 2fold
Y34F
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decreases A(-4) pre-tRNA binding affinity, although to a smaller extent than mutant Y34A. Binds A(-4) and P(-4) pre-tRNAs with comparable affinities. Mutations in the central cleft of P protein alters the observed cleavage rate constant by less than 2fold
A333C
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no effect on activity of M1 RNA on wild-type Escherichia coli SuIII tRNATyr precursor
A334U
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much lower activity on activity of M1 RNA on wild-type Escherichia coli SuIII tRNATyr precursor
G224A/G225A
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ribozymes carrying the mutation at the catalytic RNA exhibit at least 10times higher cleavage efficiency than the wild-type enzyme
N32A/E33A/K36A
mutant protein L7Ae displays only 4% of the activity observed with the wild-type protein L7Ae
H67A
spectrum of mutant is moderately altered at 37°C, indicative of subtle structural changes arising due to the mutation. The H67A protein shows significant loss in secondary structure at 45°C, which further enhances at 55°C. The loss of structure with increase in temperature is relatively less in the case of wild-type and H67N proteins. kcat/Km-value is 1.9fold lower than the value for the wild-type enzyme
H67N
spectrum of mutant is moderately altered at 37°C, indicative of subtle structural changes arising due to the mutation. The H67A protein shows significant loss in secondary structure at 45°C, which further enhances at 55°C. The loss of structure with increase in temperature is relatively less in the case of wild-type and H67N proteins. kcat/Km-value is 1.4fold lower than the value for the wild-type enzyme
H67A
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spectrum of mutant is moderately altered at 37°C, indicative of subtle structural changes arising due to the mutation. The H67A protein shows significant loss in secondary structure at 45°C, which further enhances at 55°C. The loss of structure with increase in temperature is relatively less in the case of wild-type and H67N proteins. kcat/Km-value is 1.9fold lower than the value for the wild-type enzyme
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H67N
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spectrum of mutant is moderately altered at 37°C, indicative of subtle structural changes arising due to the mutation. The H67A protein shows significant loss in secondary structure at 45°C, which further enhances at 55°C. The loss of structure with increase in temperature is relatively less in the case of wild-type and H67N proteins. kcat/Km-value is 1.4fold lower than the value for the wild-type enzyme
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A14V
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RPP21 mutant, wild-type RPP21 binds to RPP29 3fold tighter than the mutant
C71V
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the single-Cys substitutions are introduced into a Cys-less Pfu L7Ae template C71V (i.e. RNA-binding protein L7Ae, subunit of archaeal RNase P). The native C71, which is partly buried, is mutated to Val to preserve the native fold and hydrophobic core of the protein. The C71V parental reference is more active than the wild type enzyme
K42C
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mutation does not affect activity. The single-Cys substitution is introduced into a Cys-less Pfu L7Ae template C71V (i.e. RNA-binding protein L7Ae, subunit of archaeal RNase P). The native C71, which is partly buried, is mutated to Val to preserve the native fold and hydrophobic core of the protein
R46C
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mutation results in 28% decrease in activity. The single-Cys substitution is introduced into a Cys-less Pfu L7Ae template C71V (i.e. RNA-binding protein L7Ae, subunit of archaeal RNase P). The native C71, which is partly buried, is mutated to Val to preserve the native fold and hydrophobic core of the protein
V95C
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mutation results in 6% decrease in activity. The single-Cys substitution is introduced into a Cys-less Pfu L7Ae template C71V (i.e. RNA-binding protein L7Ae, subunit of archaeal RNase P). The native C71, which is partly buried, is mutated to Val to preserve the native fold and hydrophobic core of the protein
C68S/C71S
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mutant enzyme exhibits little enzymatic activity, mutation in ribonuclease P protein Ph1601p
C97S/C100S
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mutant enzyme exhibits little enzymatic activity, mutation in ribonuclease P protein Ph1601p
D180A
site-directed mutagenesis, activity of the holoenzyme reconstituted with the recombinant mutant protein subunit Ph1877p is unaltered compared to holoenzyme reconstituted with recombinant wild-type Ph1877p
D98A
site-directed mutagenesis, activity of the holoenzyme reconstituted with the recombinant mutant protein subunit Ph1877p is unaltered compared to holoenzyme reconstituted with recombinant wild-type Ph1877p
DELTAM1-R31
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RNase P reconstituted with mutant protein Ph1771p has 15% reduced activity compared to that of the reconstituted RNase P with wild-type Ph1771p
E73A
mutant shows reduced activity compared to the wild type enzyme
F95A
mutant shows reduced activity compared to the wild type enzyme
H114A
site-directed mutagenesis, activity of the holoenzyme reconstituted with the recombinant mutant protein subunit Ph1877p is unaltered compared to holoenzyme reconstituted with recombinant wild-type Ph1877p
K123A
site-directed mutagenesis, reconstitution of the holoenzyme with the recombinant mutant protein subunit Ph1877p results in reduced activity compared to holoenzyme reconstituted with recombinant wild-type Ph1877p
K158A
site-directed mutagenesis, activity of the holoenzyme reconstituted with the recombinant mutant protein subunit Ph1877p is unaltered compared to holoenzyme reconstituted with recombinant wild-type Ph1877p
K196A
site-directed mutagenesis, reconstitution of the holoenzyme with the recombinant mutant protein subunit Ph1877p results in reduced activity compared to holoenzyme reconstituted with recombinant wild-type Ph1877p
K42A
site-directed mutagenesis, activity of the holoenzyme reconstituted with the recombinant mutant protein subunit Ph1877p is unaltered compared to holoenzyme reconstituted with recombinant wild-type Ph1877p
K90A
mutant shows strongly reduced activity compared to the wild type enzyme
R105A
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mutation in ribonuclease P protein Ph1601p, mutation causes a significant reduction of the reconstituted RNase P activity as compared with that reconstituted by wild-type Ph1601p
R107A
site-directed mutagenesis, reconstitution of the holoenzyme with the recombinant mutant protein subunit Ph1877p results in reduced activity compared to holoenzyme reconstituted with recombinant wild-type Ph1877p
R115A
mutant shows reduced activity compared to the wild type enzyme
R176A
site-directed mutagenesis, reconstitution of the holoenzyme with the recombinant mutant protein subunit Ph1877p results in 78% reduced activity compared to holoenzyme reconstituted with recombinant wild-type Ph1877p
R68A
site-directed mutagenesis, reconstitution of the holoenzyme with the recombinant mutant protein subunit Ph1877p results in slightly reduced activity compared to holoenzyme reconstituted with recombinant wild-type Ph1877p
R75A
mutant shows reduced activity compared to the wild type enzyme
R87A
site-directed mutagenesis, activity of the holoenzyme reconstituted with the recombinant mutant protein subunit Ph1877p is unaltered compared to holoenzyme reconstituted with recombinant wild-type Ph1877p
R90A
site-directed mutagenesis, reconstitution of the holoenzyme with the recombinant mutant protein subunit Ph1877p results in reduced activity compared to holoenzyme reconstituted with recombinant wild-type Ph1877p
C68S/C71S
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mutant enzyme exhibits little enzymatic activity, mutation in ribonuclease P protein Ph1601p
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C97S/C100S
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mutant enzyme exhibits little enzymatic activity, mutation in ribonuclease P protein Ph1601p
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D180A
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site-directed mutagenesis, activity of the holoenzyme reconstituted with the recombinant mutant protein subunit Ph1877p is unaltered compared to holoenzyme reconstituted with recombinant wild-type Ph1877p
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DELTAM1-R31
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RNase P reconstituted with mutant protein Ph1771p has 15% reduced activity compared to that of the reconstituted RNase P with wild-type Ph1771p
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K158A
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site-directed mutagenesis, activity of the holoenzyme reconstituted with the recombinant mutant protein subunit Ph1877p is unaltered compared to holoenzyme reconstituted with recombinant wild-type Ph1877p
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R105A
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mutation in ribonuclease P protein Ph1601p, mutation causes a significant reduction of the reconstituted RNase P activity as compared with that reconstituted by wild-type Ph1601p
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R176A
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site-directed mutagenesis, reconstitution of the holoenzyme with the recombinant mutant protein subunit Ph1877p results in 78% reduced activity compared to holoenzyme reconstituted with recombinant wild-type Ph1877p
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R68A
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site-directed mutagenesis, reconstitution of the holoenzyme with the recombinant mutant protein subunit Ph1877p results in slightly reduced activity compared to holoenzyme reconstituted with recombinant wild-type Ph1877p
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R87A
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site-directed mutagenesis, activity of the holoenzyme reconstituted with the recombinant mutant protein subunit Ph1877p is unaltered compared to holoenzyme reconstituted with recombinant wild-type Ph1877p
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A347C/C348U/C353G/C354GC355AG356U
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M1-C2, is derived from M1-C1 by introducing several point mutations at the catalytic P4 domain. Is catalytically inactive
G190U/A258C
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functional ribozyme M1-C1, the 3'-terminus of an engineered M1GS ribozyme, V57, covalently linked with a guide sequence of 18 nucleotides that is complementary to the targeted mRNA sequence, which yields efficient cleavage
F17A
site-directed mutagenesis of the protein part, the mutant shows 95% reduced activity compared to the wild-type enzyme
F17A/U52C
site-directed mutagenesis of the protein part (F17A) and the RNA part (U52C), the mutant shows over 99% reduced activity compared to the wild-type enzyme
F21A
site-directed mutagenesis of the protein part, the mutant shows 80% reduced activity compared to the wild-type enzyme
K51A
site-directed mutagenesis of the protein part, the mutant shows 60% reduced activity compared to the wild-type enzyme
K53A
site-directed mutagenesis of the protein part, the mutant shows 80% reduced activity compared to the wild-type enzyme
K56A
site-directed mutagenesis of the protein part, the mutant shows 10% increased activity compared to the wild-type enzyme
K62A
site-directed mutagenesis of the protein part, the mutant shows unaltered activity compared to the wild-type enzyme
K90A
site-directed mutagenesis of the protein part, the mutant shows 70% reduced activity compared to the wild-type enzyme
R14A
site-directed mutagenesis of the protein part, the mutant shows 60% reduced activity compared to the wild-type enzyme
R15A
site-directed mutagenesis of the protein part, the mutant shows 70% reduced activity compared to the wild-type enzyme
R52A
site-directed mutagenesis of the protein part, the mutant shows 60% reduced activity compared to the wild-type enzyme
R59A
site-directed mutagenesis of the protein part, the mutant shows 20% increased activity compared to the wild-type enzyme
R60A
site-directed mutagenesis of the protein part, the mutant shows 30% reduced activity compared to the wild-type enzyme
R65A
site-directed mutagenesis of the protein part, the mutant shows 50% increased activity compared to the wild-type enzyme
R89A
site-directed mutagenesis of the protein part, the mutant shows 94% reduced activity compared to the wild-type enzyme
R89A/U52C
site-directed mutagenesis of the protein part (R89A) and the RNA part (U52C), the mutant shows over 99% reduced activity compared to the wild-type enzyme
U52C
site-directed mutagenesis of the RNA part, the mutant shows 92% reduced activity compared to the wild-type enzyme
additional information
K64C
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site-directed mutagenesis of the RNR motif of P protein subunit, the mutant shows unaltered RNA binding kinetics compared to the wild-type enzyme
K64C
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site-directed mutagenesis of an RNR motif residue, the mutant shows altered binding affinity for the substrate and between its components compared to the wild-type enzyme
R60A
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decrease in the affinity of A(-4) pre-tRNA compared to that of G(-4) pre-tRNA, causing the A(-4)/G(-4) selectivity ratio to decrease to less than 1
R60A
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site-directed mutagenesis of the RNR motif of P protein subunit, the mutant shows unaltered RNA binding kinetics compared to the wild-type enzyme
R60A
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site-directed mutagenesis of an RNR motif residue, the mutant shows altered binding affinity for the substrate and between its components compared to the wild-type enzyme
R62A
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site-directed mutagenesis of the RNR motif of P protein subunit, the mutant shows unaltered RNA binding kinetics compared to the wild-type enzyme
R62A
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site-directed mutagenesis of an RNR motif residue, the mutant shows altered binding affinity for the substrate and between its components compared to the wild-type enzyme
R65A
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affinity of RNase P for A(-4) pre-tRNA is decreased more than 10fold
R65A
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site-directed mutagenesis of the RNR motif of P protein subunit, the mutant shows unaltered RNA binding kinetics compared to the wild-type enzyme
R65A
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site-directed mutagenesis of an RNR motif residue, the mutant shows altered binding affinity for the substrate and between its components compared to the wild-type enzyme
R65C
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site-directed mutagenesis of the RNR motif of P protein subunit, the mutant shows unaltered RNA binding kinetics compared to the wild-type enzyme
R65C
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site-directed mutagenesis of an RNR motif residue, the mutant shows altered binding affinity for the substrate and between its components compared to the wild-type enzyme
R68A
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site-directed mutagenesis of the RNR motif of P protein subunit, the mutant shows unaltered RNA binding kinetics compared to the wild-type enzyme
R68A
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site-directed mutagenesis of an RNR motif residue, the mutant shows altered binding affinity for the substrate and between its components compared to the wild-type enzyme
R68C
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site-directed mutagenesis of the RNR motif of P protein subunit, the mutant shows unaltered RNA binding kinetics compared to the wild-type enzyme
R68C
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site-directed mutagenesis of an RNR motif residue, the mutant shows altered binding affinity for the substrate and between its components compared to the wild-type enzyme
additional information
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construction of mutant enzyme chimeras consisting of either Escherichia coli protein components and Bacillus subtilis RNA component or vice versa
additional information
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mutagenesis of the CR-IV region of the enzyme RPR1 RNA results in large reduction of kcat with little effect on Km
additional information
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RNR motif mutations do not alter the kinetics of pre-tRNA cleavage
additional information
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L5.1-L15.1 intradomain contact in the catalytic domain of the prototypic type B RNase P RNA of Bacillus subtilis is crucial for adopting a compact functional conformation. Disruption of the L5.1-L15.1 contact by antisense oligonucleotides or mutation reduces P RNA-alone and holoenzyme activity by one to two orders of magnitude in vitro, largely retards gel mobility of the RNA and further affects the structure of regions P7/P8/P10.1, P15 and L15.2, and abolishes the ability of Bacillus subtilis P RNA to complement a P RNA-deficient Escherichia coli strain
additional information
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construction of mutants inactive with pre-4.5S RNA but not with pre-tRNA
additional information
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construction of deletion mutants of subunit DRpp29, analysis of interaction of the mutants with RNase P subunit compared to the wild-type subunit DRpp29 and compared to the modeling data, overview
additional information
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construction of deletion mutants of subunit DRpp29, analysis of interaction of the mutants with RNase P subunit compared to the wild-type subunit DRpp29 and compared to the modeling data, overview
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additional information
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temperature-sensitive mutant that is defective in RNase P activity carries a mutant gene that has GC-AT substitutions at positions corresponding to 89 and 365 nucleotides downstream from the 5' terminus of the RNA sequence. Comparing to the wild-type RNA, the mutant RNA is less stable and rapidly degraded in vivo and vitro
additional information
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RNase P activity can be detected with holoenzymes reconstituted with the RNA subunit from Escherichia coli and the protein subunit from Synechocystis 6803 or with the RNA subunit from Synechocystis 6803 and the protein subunit from Escherichia coli
additional information
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ribozyme variant O67 contains a deletion of G336 and mutations of G224G225/A224A225, G230/C230 and A337G and exhibits 20fold higher activity to cleave the TK mRNA sequence than the wild-type ribozyme under both high and low concentrations of Mg2+ ions
additional information
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binding and catalysis of J5/15 and N(-1) mutants, overview, mutation of the A248 nucleotide results in defects in thermodynamics and in miscleavage of substrates containing 2'deoxy modification at N(-1), compensatory mutation of N(-1) restore correct cleavage activity in reaction with both, the RNA subunit alone and the holoenzyme, influence of nucleotide at position N(-2)
additional information
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construction of mutant enzyme chimeras consisting of either Escherichia coli protein components and Bacillus subtilis RNA component or vice versa
additional information
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mutagenesis of the CR-IV region of the enzyme RPR1 RNA results in large reduction of kcat with little effect on Km
additional information
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the use of Ca2+ as catalytic metal ion is enhanced in nucleotide point mutants C70U and U69 deletion in the P4 helix
additional information
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change of RNase P RNA function by single base mutation correlates with perturbation of metal ion binding in P4
additional information
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examining the bases of the J3/4 domain of Escherichia coli ribonuclease P. Replacement, deletion, or insertion, except at G63G64, can be compensated for the presence of a high concentration of magnesium ions above 20 mM
additional information
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variant with a point mutation at nucleotide 95 of RNaseP catalytic RNA G95U that increases the rate of cleavage, and another mutation at A200C that enhances substrate binding of the ribozyme
additional information
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construction of a mutant variant M1GS ribozyme via combined mutations at nucleotide 83 and 340 of RNase P catalytic RNA (G83 -. U83 and G340 -. A340). The mutant ribozyme shows an increase in overall efficiency in cleaving an HIV RNA sequence compared to the wild-type enzyme
additional information
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construction of a rnpA knockout strain (MTea1), deletion of the essential endogenous rnpA copy and simultaneous replacement by a heterologous version of the gene, i.e. by eight rnpA sequences from Proteus mirabilis, Pseudomonas aeruginosa, Acinetobacter baumannii, Neisseria gonorrhoaea, Bacillus subtilis, Streptococcus oralis, Staphylococcus aureus, and Thermotoga maritima. The increasingly divergent versions of the RNase P protein all complement the loss of the endogenous rnpA gene, phylogenetic range of rnpA heterologues, overview
additional information
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construction of the enzyme-deficient rnpA49 rph-1 mutant strain. Increased levels of the M1 RNA, the enzyme's rNA subunit, partially complement the rnpA49 allele at 42°C. Simultaneous expression of tRNAval(GAC) and tRNAval(UAC) does not suppress the temperature sensitivity of the rnpA49 strain. In contrast, overproduction of the rnpB gene leads to a small improvement in the growth of the rnpA49 rph-1 strain at 42°C
additional information
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seperation of the catalytic domain of the RNA part by cleaving it from the specificity S-domain mediates. Compared to full-length RNase P RNA, the rate constant, kobs, of cleavage of various model RNA hairpin loop substrates.for the truncated RPR is reduced 30 to 13000fold depending on the substrate. Substitution of A248, positioned near the cleavage site in the RNase P-substrate complex, with G in the truncated RNase P RNA results in 30fold improvement in reaction rate. In contrast, strengthening the interaction between the RNase P RNA and the 3'-end of the substrate only had a modest effect. Deleting the S-domain gives a reduction in the rate, but it results in a less erroneous RPR with respect to cleavage site selection
additional information
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construction of the enzyme-deficient rnpA49 rph-1 mutant strain. Increased levels of the M1 RNA, the enzyme's rNA subunit, partially complement the rnpA49 allele at 42°C. Simultaneous expression of tRNAval(GAC) and tRNAval(UAC) does not suppress the temperature sensitivity of the rnpA49 strain. In contrast, overproduction of the rnpB gene leads to a small improvement in the growth of the rnpA49 rph-1 strain at 42°C
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additional information
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generation of several enzyme mutants with deleted stem loops in their RNA part. The mutants show reduced activity
additional information
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in order to investigate their functional role, six mutants are propared, DELTAP1, DELTAP3, DELTAP8, DELTAP9, DELTAP12, and DELTAP15, in which the stem-loops including helices P1, P3, P8, P9, P12/12.1/12.2, and P15/16 are individually deleted respectively. the mutant proteins are characterized with respect to pre-tRNA cleavage activity in the presence of five proteins and also to the ability to form a complex with the proteins. The results indicate that elimination of the stem-loops results in a reduction of the pre-tRNA cleavage activity. It is further suggested that the stem-loops containing P3 or P15/16 form a binding site for the PhoPop5-PhoRpp30 complex, and that their interaction is closely involved in the structural formation of an active site in PhopRNA
additional information
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generation of several enzyme mutants with deleted stem loops in their RNA part. The mutants show reduced activity
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additional information
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in order to investigate their functional role, six mutants are propared, DELTAP1, DELTAP3, DELTAP8, DELTAP9, DELTAP12, and DELTAP15, in which the stem-loops including helices P1, P3, P8, P9, P12/12.1/12.2, and P15/16 are individually deleted respectively. the mutant proteins are characterized with respect to pre-tRNA cleavage activity in the presence of five proteins and also to the ability to form a complex with the proteins. The results indicate that elimination of the stem-loops results in a reduction of the pre-tRNA cleavage activity. It is further suggested that the stem-loops containing P3 or P15/16 form a binding site for the PhoPop5-PhoRpp30 complex, and that their interaction is closely involved in the structural formation of an active site in PhopRNA
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additional information
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complete deletion of the protein subunit is lethal, mutation of P3 loop causes defects in pre-tRNA processing and enzyme RPR1 RNA maturation as well as subunit interaction in vivo
additional information
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construction of a pre-tRNA substrate in which the pro-Rp nonbridging oxygen of the scissile bond is replaced by sulfur
additional information
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mutations in yeast RPR1 RNA at the loop regions of eP8 hairpin (eP8m), eP9 hairpin (eP9m), the junction of P4/P7 (jP4/7m), and the junction of P7/eP15 (jP7/15m). The mutation of jP7/15m does not affect cell growth, tRNA processing, or RNase P maturation. Mutants in eP8m, eP9m, and jP4/7m display pronounced growt defects and pre-tRNA processing defects in vivo
additional information
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Trypanosoma brucei proteinaceous enzyme PRORP1 can substitute for yeast nuclear RNase P in vivo
additional information
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construction of a functional RNase P-based ribozyme, M1GS RNA, that targets the overlapping mRNA region of M80.5 and protease, two murine cytomegalovirus proteins essential for viral replication. Construction of an attenuated strain of Salmonella, which exhibits efficient gene transfer activity and little cytotoxicity and pathogenicity in mice, for delivery of anti-MCMV ribozyme. In MCMV-infected macrophages treated with the constructed attenuated Salmonella strain carrying the functional M1GS RNA construct, a 80-85% reduction in the expression of M80.5/protease and a 2500fold reduction in viral growth is observed. The strain delivers efficiently delivered antiviral M1GS RNA into spleens and livers, leading to substantial expression of the ribozyme without causing significant adverse effects in the animals. MCMV-infected mice treated orally with Salmonella carrying the functional M1GS sequence display reduced viral gene expression, decreased viral titers, and improved survival compared to the untreated mice or mice treated with Salmonella containing control ribozyme sequences
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RNase P activity can be detected with holoenzymes reconstituted with the RNA subunit from Escherichia coli and the protein subunit from Synechocystis 6803 or with the RNA subunit from Synechocystis 6803 and the protein subunit from Escherichia coli
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wild-type and mutant enzyme structure-function analysis, overview. Point mutations in the conserved protein loop (residues 52-57) have either no or modest effects on catalytic efficiency. Similarly, amino acid changes in the RNR region, which represent the most conserved region of bacterial RNase P proteins, exhibit negligible changes in catalytic efficiency
additional information
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wild-type and mutant enzyme structure-function analysis, overview. Point mutations in the conserved protein loop (residues 52-57) have either no or modest effects on catalytic efficiency. Similarly, amino acid changes in the RNR region, which represent the most conserved region of bacterial RNase P proteins, exhibit negligible changes in catalytic efficiency
