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Literature summary for 3.1.26.11 extracted from

  • Karkashon, S.; Hopkinson, A.; Levinger, L.
    tRNase Z catalysis and conserved residues on the carboxy side of the His cluster (2007), Biochemistry, 46, 9380-9387.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

Cloned (Comment) Organism
baculovirus expression system, pFastbac HTA, transformation into insect cells Drosophila melanogaster

Protein Variants

Protein Variants Comment Organism
A631T HEAT-domain-variant Drosophila melanogaster
C584A motif I-variant Drosophila melanogaster
D610A motif IV-variant Drosophila melanogaster
E630A HEAT-domain-variant Drosophila melanogaster
F669A motif V-variant Drosophila melanogaster
G609A motif IV-variant Drosophila melanogaster
H583A motif I-variant Drosophila melanogaster
H629A HEAT-domain-variant Drosophila melanogaster
H646A HST loop-variant Drosophila melanogaster
H668A motif V-variant Drosophila melanogaster
K644A HST loop-variant Drosophila melanogaster
L643A HST loop-variant Drosophila melanogaster
M612A motif IV-variant Drosophila melanogaster
additional information Ala-scanning mutagenesis through five conserved loops on the carboxy side of motif II (motifs III, IV, HEAT,HST, and motif V) performed, pre-tRNA processing kinetics of the expressed variants studied Drosophila melanogaster
N586A motif I-variant Drosophila melanogaster
P585A motif I-variant Drosophila melanogaster
P613A motif IV-variant Drosophila melanogaster
Q651A HST loop-variant Drosophila melanogaster
Q671A motif V-variant Drosophila melanogaster
R582A motif I-variant Drosophila melanogaster
R642A HST loop-variant Drosophila melanogaster
R672A motif V-variant Drosophila melanogaster
S587A motif I-variant Drosophila melanogaster
S608A motif IV-variant Drosophila melanogaster
S647A HST loop-variant Drosophila melanogaster
S650A HST loop-variant Drosophila melanogaster
S670A motif V-variant Drosophila melanogaster
T611A motif IV-variant Drosophila melanogaster
T632A HEAT-domain-variant Drosophila melanogaster
T645A HST loop-variant Drosophila melanogaster
T648A HST loop-variant Drosophila melanogaster
T667A motif V-variant Drosophila melanogaster
V581A motif I-variant Drosophila melanogaster
V649A HST loop-variant Drosophila melanogaster
Y607A motif IV-variant Drosophila melanogaster
Y673A motif V-variant Drosophila melanogaster

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
additional information kinetics of tRNase Z processing in wild-type and variants shown, effects of substitutions in conserved regions on the C-side of motif II indicated, KM-values for tested variants similar to the mean wild-type KM value of 15 nM, residues involved with metal ion binding and catalysis not directly involved with substrate recognition and binding Drosophila melanogaster

Metals/Ions

Metals/Ions Comment Organism Structure
Zn2+ binding of two zinc ions coordinated by four residues in the signature His domain, catalytically important conserved contact between Glu231 and His24 shown Drosophila melanogaster

Organism

Organism UniProt Comment Textmining
Drosophila melanogaster Q8MKW7
-
-

Purification (Commentary)

Purification (Comment) Organism
nickel chelate affinity purification Drosophila melanogaster

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
additional information
-
reaction kinetics of tRNase Z variants with Ala substitutions through conserved loops on the carboxy side of motif II shown, substitution of motif IV aspartate reduces catalytic efficiency more than 10000fold, Histidines in motif III, V, and the HST loop also functionally important, replacement of Glu in the HEAT loop with Ala reduces efficiency by about 1000fold Drosophila melanogaster

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
pre-tRNAarg + H2O the new substrate pre-tRNAArg reveals kinetic parameters with wild type tRNase Z similar to those reported for pre-tRNAHis, enzyme concentration of 25 pM Drosophila melanogaster ?
-
?

Synonyms

Synonyms Comment Organism
tRNase Z
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Drosophila melanogaster