Please wait a moment until all data is loaded. This message will disappear when all data is loaded.
Please wait a moment until the data is sorted. This message will disappear when the data is sorted.
K159M
site-directed mutagenesis, the mutation changes the rate-limiting step to the hydride transfer, proton transfer is blocked in the mutant but can be rescued using exogenous proton acceptors, such as buffers, small primary amines, and azide, overview
P185A
site-directed mutagenesis, analysis of kinetics and structure compared to the wild-type enzyme
P185G
site-directed mutagenesis, analysis of kinetics and structure compared to the wild-type enzyme
S114A/Y155F
-
site-directed mutagenesis, the mutant shows altered kinetics and pH profile, and 200000fold reduced activity compared to the wild-type
T188A
site-directed mutagenesis, analysis of kinetics and structure compared to the wild-type enzyme
T188S
site-directed mutagenesis, analysis of kinetics and structure compared to the wild-type enzyme
W173F/P185W
site-directed mutagenesis, analysis of kinetics and structure compared to the wild-type enzyme
W173F/T188W
site-directed mutagenesis, analysis of kinetics and structure compared to the wild-type enzyme
Y155F/K159A
-
site-directed mutagenesis, the mutant shows altered kinetics and pH profile, and 9400fold reduced activity compared to the wild-type
V54L
site-directed mutagenesis, the change renders the sequence identical to that of human 20-alpha hydroxysteroid dehydrogenase. The V54L mutation directly restricts the steroid binding modes to a unique one, which resembles the orientation of 20alpha-OHProg within human 20alpha-HSD. The kinetic study shows that the V54L mutation significantly decreases the 3alpha-HSD activity for the reduction of 5alpha-dihydrotestosterone, while this mutation enhances the 20alpha-HSD activity to convert progesterone
K157A
the NADH binding affinity of K157A mutant is much lower than that of the wild-type, mainly due to loss of a hydrogen bond
S114A
mutant shows higher Km and lower kcat values in both oxidation and reduction reactions
S114A/Y153F
double mutation results in a significant decrease in kcat relative to the single mutant Y153F
Y153F
mutant shows higher Km and lower kcat values in both oxidation and reduction reactions. Loss of hydrogen bonding with NADH upon the Y153F mutation results in increased enthalpy change
E276R
-
site-directed mutagenesis, the mutation alters the cofactor specificity of AKR1C17 from NAD+ to NADP+, the switch is analogy th the residues of AKR1C9 and its cofactor specificity, overview
F129A
-
site-directed mutagenesis, mutation of a substrate binding residue, altered steroid recognition and kinetics compared to the wild-type enzyme, highly reduced activity
Q270K
-
site-directed mutagenesis, the mutation alters the cofactor specificity of AKR1C17 from NAD+ to NADP+, the switch is analogy th the residues of AKR1C9 and its cofactor specificity, overview
Q270K/E276R
-
site-directed mutagenesis, the mutation alters the cofactor specificity of AKR1C17 from NAD+ to NADP+, the switch is analogy th the residues of AKR1C9 and its cofactor specificity, overview
R276E
-
site-directed mutagenesis, the mutant shows increased preference for the oxidation reaction compared to the wild-type enzyme
R276G
-
site-directed mutagenesis, the mutant shows slightly increased preference for the reduction reaction compared to the wild-type enzyme
R276M
-
site-directed mutagenesis, the mutant shows slightly increased preference for the reduction reaction compared to the wild-type enzyme
D249A
the mutation leads to 2fold increased Km value compared to the wild type, the mutant shows increased retention time, suggesting a smaller molecule size than dimeric wild type enzyme
D249A
mutation interrupts salt bridge between residues D249 and R167, secondary structure similar to wild-type. 30fold decrease in catalytic efficiency, decrease in melting temperature
D249K
he mutation leads to 4fold increased Km value compared to the wild type, the mutant shows increased retention time, suggesting a smaller molecule size than dimeric wild type enzyme
D249K
mutation interrupts salt bridge between residues D249 and R167, secondary structure similar to wild-type. 1.4fold decrease in catalytic efficiency, decrease in melting temperature
D249S
the mutant has similar kinetic parameters to wild type enzyme
D249S
mutation interrupts salt bridge between residues D249 and R167, secondary structure similar to wild-type. 1400fold decrease in catalytic efficiency, decrease in melting temperature
K159A
-
site-directed mutagenesis, the mutant shows altered kinetics and pH profile, and 200fold reduced activity compared to the wild-type
K159A
site-directed mutagenesis, the mutation changes the rate-limiting step to the hydride transfer, proton transfer is blocked in the mutant but can be rescued using exogenous proton acceptors, such as buffers, small primary amines, and azide, overview
K159A
decrease in the catalytic constant by 56fold and increase in the dissociation constant by 75fold. The enzyme-bound NADH decreases the fluorescence anisotropy value in the decreasing order WT, N86A, Y155F, K159A, indicating an increase in the mobility of the bound NADH for the mutants. Hydrogen bonding with the hydroxyl group of nicotinamide ribose by K159 and Y155 is important to maintain the orientation of NADH and contributes greatly to the transition-state binding energy to facilitate the catalysis
K159A
decrease in catalytic constant and increase in the dissociation constant. The enzyme-bound NADH decreases the fluorescence anisotropy value in the decreasing order WT, N86A, Y155F, K159A, indicating an increase in the mobility of the bound NADH for the mutants. Hydrogen bonding with the hydroxyl group of nicotinamide ribose by residues K159 and Y155 is important to maintain the orientation of NADH and contributes greatly to the transition-state binding energy to facilitate the catalysis. Residue N86 is important for stabilizing the position of K159
N86A
decrease in the catalytic constant by 37fold and increase in the dissociation constant by 8fold. The enzyme-bound NADH decreases the fluorescence anisotropy value in the decreasing order WT, N86A, Y155F, K159A, indicating an increase in the mobility of the bound NADH for the mutants. Residue N86 is important for stabilizing the position of K159
N86A
decrease in catalytic constant and increase in the dissociation constant. The enzyme-bound NADH decreases the fluorescence anisotropy value in the decreasing order WT, N86A, Y155F, K159A, indicating an increase in the mobility of the bound NADH for the mutants. Hydrogen bonding with the hydroxyl group of nicotinamide ribose by residues K159 and Y155 is important to maintain the orientation of NADH and contributes greatly to the transition-state binding energy to facilitate the catalysis. Residue N86 is important for stabilizing the position of K159
S114A
-
site-directed mutagenesis, the mutant shows altered kinetics and pH profile, and 3400fold reduced activity compared to the wild-type
S114A
the mutation eliminates the hydrogen bonding interaction with P185, causing a conformational change in a nonproductive binding of NADH and a significant loss of activity, the mutant enzyme decreases 3100fold in V/Et value with no apparent change in Km value for substrates
S114A
mutant enzyme exhibits a pronounced increase in the magnitude of ellipticity at 222 nm. S114A mutant enzyme decreases 3100fold in catalytic efficiency with no apparent change in Km for substrates. Addition of NADH to S114A mutant enzyme induces a secondary structural change
Y155F
-
site-directed mutagenesis, the mutant shows altered kinetics and pH profile, and 2800fold reduced activity compared to the wild-type
Y155F
decrease in the catalytic constant by 220fold and increase in the dissociation constant by 3fold. The enzyme-bound NADH decreases the fluorescence anisotropy value in the decreasing order WT, N86A, Y155F, K159A, indicating an increase in the mobility of the bound NADH for the mutants. Hydrogen bonding with the hydroxyl group of nicotinamide ribose by K159 and Y155 is important to maintain the orientation of NADH and contributes greatly to the transition-state binding energy to facilitate the catalysis
Y155F
decrease in catalytic constant and increase in the dissociation constant. The enzyme-bound NADH decreases the fluorescence anisotropy value in the decreasing order WT, N86A, Y155F, K159A, indicating an increase in the mobility of the bound NADH for the mutants
F118A
-
site-directed mutagenesis, altered kinetic and catalytic efficiency compared to the wild-type enzyme
F118A
-
site-directed mutagenesis, mutation of a substrate binding residue, altered steroid recognition and kinetics compared to the wild-type enzyme
L54A
-
site-directed mutagenesis, altered kinetic and catalytic efficiency compared to the wild-type enzyme
L54A
-
site-directed mutagenesis, mutation of a substrate binding residue, altered steroid recognition and kinetics compared to the wild-type enzyme
N306A
-
site-directed mutagenesis, altered kinetic and catalytic efficiency compared to the wild-type enzyme
N306A
-
site-directed mutagenesis, mutation of a substrate binding residue, altered steroid recognition and kinetics compared to the wild-type enzyme
T226A
-
site-directed mutagenesis, altered kinetic and catalytic efficiency compared to the wild-type enzyme
T226A
-
site-directed mutagenesis, mutation of a substrate binding residue, steroid recognition and kinetics are similar to the wild-type enzyme
T24A
-
site-directed mutagenesis, altered kinetic and catalytic efficiency compared to the wild-type enzyme
T24A
-
site-directed mutagenesis, mutation of a substrate binding residue, steroid recognition and kinetics are similar to the wild-type enzyme
W227A
-
site-directed mutagenesis, altered kinetic and catalytic efficiency compared to the wild-type enzyme
W227A
-
site-directed mutagenesis, mutation of a substrate binding residue, altered steroid recognition and kinetics compared to the wild-type enzyme, highly reduced activity and binding of progesterone
Y310A
-
site-directed mutagenesis, altered kinetic and catalytic efficiency compared to the wild-type enzyme
Y310A
-
site-directed mutagenesis, mutation of a substrate binding residue, altered steroid recognition and kinetics compared to the wild-type enzyme
additional information
construction of insertion mutants, overview
additional information
-
recombinant AKR1C2 and AKR1C3 mediated prostaglandin D2 metabolism augments the PI3K/Akt proliferative signaling pathway in transfected human prostate cancer cells, overview
additional information
enzyme silencing by specific siRNA suppresses 3alpha-HSD3 expression without interfering with 3alpha-HSD4, which shares a highly homologous active site, the 5alpha-DHT concentration increases, whereas MCF7 cell growth is suppressed