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1.17.4.1: ribonucleoside-diphosphate reductase

This is an abbreviated version!
For detailed information about ribonucleoside-diphosphate reductase, go to the full flat file.

Word Map on EC 1.17.4.1

Reaction

2'-deoxyribonucleoside 5'-diphosphate
+
thioredoxin disulfide
 +
H2O
=
ribonucleoside 5'-diphosphate
 +
thioredoxin

Synonyms

2'-deoxyribonucleoside-diphosphate:oxidized-thioredoxin 2'-oxidoreductase, ADP reductase, CDP reductase, class I ribonucleotide reductase, class I ribonulceotide reductase, class I RNR, class I RR, class Ia ribonucleotide reductase, class Ia RNR, class Ia RR, class Ib ribonucleotide reductase, class Ib RNR, class Ic ribonucleotide reductase, class II RNR, manganese-ribonucleotide reductase, Mn-RNR, More, mRR, NrdA, NrdB, NrdE, NrdF, nucleoside diphosphate reductase, p53-inducible ribonucleotide reductase, R2F, reductase, ribonucleoside diphosphate, ribonucleoside 5'-diphosphate reductase, ribonucleoside diphosphate reductase, ribonucleoside-diphosphate reductase subunit M2 B, ribonucleotide diphosphate reductase, ribonucleotide reductase, RIR1, RIR2, RNR, RNR1 rRibonucleoside-diphosphate reductase large chain 1, RNR2, UDP reductase

ECTree

     1 Oxidoreductases
         1.17 Acting on CH or CH2 groups
             1.17.4 With a disulfide as acceptor
                1.17.4.1 ribonucleoside-diphosphate reductase

Engineering

Engineering on EC 1.17.4.1 - ribonucleoside-diphosphate reductase

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PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
morme
AtRNR2B induction is abolished in the rad9-rad17 double mutant, transgenic plant phenotypes, overview
F127Y
-
similar CDP reductase activity as wild-type
F127Y/Y129F
-
10-15% of wild-type CDP reductase activity
W51F
-
site-directed mutagenesis, the decay of the Mn(IV)/Fe(IV) intermediate is slightly affected
Y129F
-
no CDP reductase activity
Y222F
-
the substitution by site-directed mutagenesis retards the intrinsic decay of the Mn(IV)/Fe(IV) intermediate by about 10fold and diminishes the ability of ascorbate to accelerate the decay by about 65fold but has no detectable effect on the catalytic activity of the Mn(IV)/Fe(III)-R2 product
Y338F
-
site-directed mutagenesis, substitution of Y338, the cognate of the subunit interfacial R2 residue in the R1 S R2 PCET pathway of the conventional class I RNRs, has almost no effect on decay of the Mn(IV)/Fe(IV) intermediate but abolishes catalytic activity
G392S
-
temperature-sensitive protein with complete splicing activity at 17 and 28°C but not at 37°C or higher
G392S/C539G
-
the cleavage at the ribonucleotide reductase RIR1 intein C-terminus is blocked, but other cleavage activities can be efficiently performed at 17°C. The mutant variant possesses the properties of low-temperature-induced cleavage at the intein N-terminus
C225A
-
4-6% of wild-type activity
C225S
C439A
-
4-6% of wild-type activity
C439S
-
the C439S mutant of the Escherichia coli R1 is catalytically inactive in vitro
C462S
-
in the presence of dithiothreitol the major product formed by interaction with CDP is cytosine
C754A
-
active with dithiothreitol as reductant, 3% of wild type activity with thioredoxin
C759A
-
active with dithiothreitol as reductant, 3% of wild type activity with thioredoxin
C759S
-
C759 may play a role in the relay of electrons between thioredoxin and subunit B1
D84E
the mutation provides a ligand environment similar to that found in methane monooxygenase, MMO, renders this residue bidentate, and Glu204 becomes monodentate. The RNR mutant, however, remains distinct from MMO, which has a beta-1,1 type Glu-bridge, most likely due to the effects of second sphere residues
N238A
-
the monomeric R1 protein is able to dimerize when bound by both substrate and effector and is able to reduce ribonucleotides with a comparatively high capacity
W48A/Y122F
the reaction remains at the level of the peroxo-intermediate, structural analysis
W48F/D84E
the reaction remains at the level of the peroxo-intermediate, structural analysis
Y122F
Y122F/Y356F
Y122H
the specific activity of mutant enzyme preparation is less than 0.5% of the wild-type activity. Mutant of R2 protein subunit, the mutant contains a novel stable paramagnetic center, named H. Deteiled characterization of center H, using 1H2H-14N/15N- and 57Fe-EDDOR in comparison with the FeIIIFeIV intermediate X observed in the iron reconstitution reaction of R2, a new tyrosyl radical on Phe208 as ligand to the diiron center
Y356F
Y730F
-
site-directed mutagenesis, the mutant excludes a direct superexchange mechanism between C439 and Y731 in radical transport, overview
D16R
-
site-directed mutagenesis, the mutant retains 55% of wild-type activity for CDP reduction, and 67% for ADP reduction, it is not inhibited and does not form hexamers at physiologically relevant dATP concentrations
D287A
H2E
-
site-directed mutagenesis, the mutant retains 56% of wild-type activity for CDP reduction, and 56% for ADP reduction
K95E
mutation in small subunit M2, results in dimer disassembly and enzyme activity inhibition. Mutant is capable of generating the diiron and tyrosyl radical cofactor, but the disassembly of the M2 dimer reduces its interaction with the large subunit M1. The transfection of the wild-type M2 but not the K95E mutant rescues theG1/S phase cell cycle arrest and cell growth inhibition caused by the siRNA knockdown of M2
K95E/E98K
charge-exchanging double mutation, recovers the dimerization and activity lost in mutant K95E
R265A
-
mutant in subunit R2, about 10% of wild-type activity. Mutant is able to form stable tyrosyl radicals and bind subunit R1 with similar kinetics as wild-type
R265E
-
mutant in subunit R2, about 40% of wild-type activity. Mutant is able to form stable tyrosyl radicals and bind subunit R1 with similar kinetics as wild-type
R265Q
-
mutant in subunit R2, about 1% of wild-type activity. Mutant is able to form stable tyrosyl radicals and bind subunit R1 with similar kinetics as wild-type
R265Y
-
mutant in subunit R2, about 4% of wild-type activity. Mutant is able to form stable tyrosyl radicals and bind subunit R1 with similar kinetics as wild-type
Y177F
-
tyrosyl residue involved in radical formation
Y370F
-
mutation in R2 subunit, no activity
Y370W
-
mutation in R2 subunit, point mutation does not affect the ability to form a normal diferric iron/tyrosyl radical center, 1.7% of wild-type activity probably due to slow radical transfer
E106A/E126A
mutant enzyme completely loses the ability to be inhibited by dATP. Like the wild-type protein the mutant enzyme can bind approximately three dATP per polypeptide. The mutant enzyme loses the ability to tetramerize and only forms dimers regardless of allosteric effector
H72A/D73A/Y830A
the mutant enzyme forms inactive tetramers in the presence of any allosteric effector, the mutant enzyme partially loses its propensity to be inhibited by dATP. Like the wild-type protein the mutant enzyme can bind approximately three dATP per polypeptide
R119D
mutant enzyme completely loses the ability to be inhibited by dATP. Like the wild-type protein the mutant enzyme can bind approximately three dATP per polypeptide. The mutant enzyme loses the ability to tetramerize and only forms dimers regardless of allosteric effector
C428S
-
mutantion is lethal. Cells carrying both the C428S and the SX2S mutation of CX2C motif on plasmids are viable and form colonies with an efficiency similar to that of the wild-type control showing interallelic complementation
K387N
-
affords higher activity due to increased tyrosyl radical content
Q288A
mutation causes severe S phase defects in cells that use the enzyme as the sole source of of ribonucleoside diphosphate activity. Compared to the wild-type enzyme activity, Q288A mutants show 15% of ADP reduction, whereas they show 23% of CDP reduction. There is a 6fold loss of affinity for ADP binding and a 2fold loss of affinity for CDP. Q288A can support mitotic growth, albeit with a severe S phase defect
R293A
mutation causes lethality in cells that use the enzyme as the sole source of ribonucleoside diphosphate activity. Compared to the wild-type enzyme activity, R293A mutants show 4% of ADP reduction, whereas they show 20% CDP reduction. The mutant is unable to bind ADP and binds CDP with 2fold lower affinity compared to wild-type. X-ray structures of R293A complexed with dGTP and AMPPNP reveal that ADP is not bound at the catalytic site, and CDP binds farther from the catalytic site compared to wild type. R293A cannot support mitotic growth
additional information