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A96L/N212K
mutation increases Km value and eliminates substrate inhibition
I283V |
site-directed mutagenesis, the mutation switches the nature of the residue from Amblyrhynchus cristatus to that of Iguana iguana, another Galapagos marine iguana, the mutation does not affect enzyme kinetics, overview
T9A
site-directed mutagenesis, the mutation switches the nature of the residue from Amblyrhynchus cristatus to that of Iguana iguana, another Galapagos marine iguana, the mutation does not affect enzyme kinetics, overview
T9A/I283V
site-directed mutagenesis, the mutations switch the nature of the residues from Amblyrhynchus cristatus to those of Iguana iguana, another Galapagos marine iguana, the mutation affects enzyme kinetics decreasing Km for pyruvate and kcat, overview
H171C
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site-directed mutagenesis
C210S
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crystallization of mutant enzyme
D241N
site-directed mutagenesis, the mutant hsows higher activity at pH above 5.5 compared to the wild-type enzyme
E60Q
site-directed mutagenesis, the mutant has a similar pH prodile as the wild-type
D241N
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site-directed mutagenesis, the mutant hsows higher activity at pH above 5.5 compared to the wild-type enzyme
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E60Q
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site-directed mutagenesis, the mutant has a similar pH prodile as the wild-type
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H88X/H226X
substitution of His 88 and 226 of the eLDHA monomer alters the surface charge of equine LDH tetramer, the residues are located in an important region affecting the catalytic kinetics
D38E
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site-directed mutagenesis, the mutant shows a twofold reduced substrate inhibition by pyruvate compared to the wild-type enzyme
D38R
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site-directed mutagenesis, the mutant shows a threefold reduced substrate inhibition by pyruvate compared to the wild-type enzyme
F16Q/C81S/N85R
catalytic efficiency is higher than that of wild-type enzyme, utilizes NAD+ better than wild-type enzyme, weakly active wth NADP+
F16Q/I37K/D38S/C81S/N85R
utilizes NADP+ better than wild-type enzyme, prefers NADP+ to NAD+
S100M
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a hybrid gene is constructed from fragments of the LDH genes from Bacillus stearothermophilus (coding for aa 15-100) and Bacillus megaterium (coding for aa 101-331). The hybrid LDH, named S100M, is more thermostable than Bacillus megaterium LDH, less thermostable than Bacillus stearothermophilus LDH and unlike the two wild-type enzymes, it can not be activated by D-fructose 1,6-bisphosphate
I229A
30% decrease in Km value and 2.6fold increase in kcat value for phenylpyruvate
Q88R
17% decrease in Km value and 3fold increase in kcat value for phenylpyruvate
T235G
slight decrease in specific activity towards phenylpyruvate
N197D
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1.15fold increase in activity, 2.7fold increase in kcat/Km ratio, 2fold increase in production titer for asymmetric synthesis of (S)-2-hydroxybutanoic acid
S163L
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decrease in substrate inhibition, the Km value for pyruvate is increased substantially and the turnover number is 60% that of wild type
S100M
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a hybrid gene is constructed from fragments of the LDH genes from Bacillus stearothermophilus (coding for aa 15-100) and Bacillus megaterium (coding for aa 101-331). The hybrid LDH, named S100M, is more thermostable than Bacillus megaterium LDH, less thermostable than Bacillus stearothermophilus LDH and unlike the two wild-type enzymes, it can not be activated by D-fructose 1,6-bisphosphate
D8G
no measurable effect on the Km value for pyruvate, increased thermal stability
R157L
mutation appears to deregulate activation by fructose 1,6-bisphosphate, leading to constitutive activation of lactate dehydrogenase
R157L
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mutation appears to deregulate activation by fructose 1,6-bisphosphate, leading to constitutive activation of lactate dehydrogenase
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additional information
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construction of an insertion mutant of gene AA02769, inactivation of AA02769 eliminates the ability of the organism to grow on L-lactate, but not on D-glucose
additional information
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construction of an insertion mutant of gene AA02769, inactivation of AA02769 eliminates the ability of the organism to grow on L-lactate, but not on D-glucose
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additional information
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construct an ethanologenic Bacillus subtilis strain, the ldh gene is disrupted in strain WB700 by chromosomal insertion of the Zymomonas mobilis pyruvate decarboxylase gene pdc and the alcohol dehydrogenase II gene adhB under the control of the ldh native promoter leading to production of etahnol and butanediol by the recombinant strain. But cell growth and glucose consumption rates in strain BS35 are reduced by 70 and 65%, respectively, in comparison to the progenitor strain. To eliminate butanediol production, the acetolactate synthase gene alsS is inactivated resulting in BS36, additional expression of transhydrogenase encoded by gene udhA from Escherichia coli allows a partial recovery of the cell growth rate and an early onset of ethanol production resulting in strain BS37, overview
additional information
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construct an ethanologenic Bacillus subtilis strain, the ldh gene is disrupted in strain WB700 by chromosomal insertion of the Zymomonas mobilis pyruvate decarboxylase gene pdc and the alcohol dehydrogenase II gene adhB under the control of the ldh native promoter leading to production of etahnol and butanediol by the recombinant strain. But cell growth and glucose consumption rates in strain BS35 are reduced by 70 and 65%, respectively, in comparison to the progenitor strain. To eliminate butanediol production, the acetolactate synthase gene alsS is inactivated resulting in BS36, additional expression of transhydrogenase encoded by gene udhA from Escherichia coli allows a partial recovery of the cell growth rate and an early onset of ethanol production resulting in strain BS37, overview
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additional information
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transposon mutagenesis of a reporter strain carrying a chromosomal ldhA promoter-lacZ fusion, PldhA-lacZ, using Tn5-based minitransposon system reveals that ldhA disruption drastically decreases expression of PldhA-lacZ. ldhA Promoter activity is reduced in the ldhA mutant. PldhA-lacZ expression in the ldhA mutant is restored by deletion of lldR, suggesting that LldR acts as a repressor of ldhA in the absence of L-lactate and the LldR-mediated repression is not relieved in the ldhA mutant due to its inability to produce L-lactate. lldR deletion does not affect PldhA-lacZ expression in the wild-type background during growth on either glucose, acetate, or L-lactate. However, it upregulates PldhA-lacZ expression in the sugR mutant background during growth on acetate. The binding sites of LldR and SugR are located around the -35 and -10 regions of the ldhA promoter, respectively
additional information
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construction of ldh1 and ldh-2 deletion mutants and of a catalytically inactive double knockout mutant of genes ldh-1 and ldh-2 by gene replacement achieved by double-crossover homologous recombination. The ldh-2 mutant shows hardly any differences in growth and metabolism in comparison to the wild-type. The two mutants lacking ldh-1, on the other hand, produce much less lactate than the wild-type but grew to higher cell densities and have higher final pHs, phenotypes, overview
additional information
deletion of Glu 14 of the eLDHB monomer alters the surface charge of equine LDH tetramers and the residue is located in an important region affecting the catalytic kinetics
additional information
deletion of Glu 14 of the eLDHB monomer alters the surface charge of equine LDH tetramers and the residue is located in an important region affecting the catalytic kinetics
additional information
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deletion of Glu 14 of the eLDHB monomer alters the surface charge of equine LDH tetramers and the residue is located in an important region affecting the catalytic kinetics
additional information
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the malate dehydrogenase, EC 1.1.1.37, mutant I12V/R81Q/M85E/G210A/V214I shows a substrate specificity that is switched from malate dehydrogenase to that of lactate dehydrogenase, overview
additional information
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a mutant into which an additional loop has been engineered in order to prevent tetramerization
additional information
the malate dehydrogenase, EC 1.1.1.37, mutant I12V/R81Q/M85E/G210A/V214I shows a substrate specificity that is switched from malate dehydrogenase to that of lactate dehydrogenase, overview
additional information
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knockdown of LDH-A by shRNA
additional information
silencing of LDHB with a small interfering RNA (siRNA) (siLDHB-2) decreasing cell number in all the cancer cell lines investigated
additional information
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construction of a Pichia stipitis strain CBS6054, that expresses the LDH from Lactobacillus helveticus under the control of the Pichia stipitis fermentative ADH1 promoter, the mutant yeast uses xylose, glucose, or a mixture of the two sugars as the carbon source for lactate production, LDH competes efficiently with the ethanol pathway for pyruvate, overview
additional information
construction of an ldh gene insertion mutant strain FI9078, the insertion of an IS905-like element, that created a hybrid promoter in the intergenic region upstream of ldhB, leads to activation of a second isozyme LDHB, which shows a strongly pH-dependent activity, overview
additional information
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construction of an ldh gene insertion mutant strain FI9078, the insertion of an IS905-like element, that created a hybrid promoter in the intergenic region upstream of ldhB, leads to activation of a second isozyme LDHB, which shows a strongly pH-dependent activity, overview
additional information
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construction of an ldh gene insertion mutant strain FI9078, the insertion of an IS905-like element, that created a hybrid promoter in the intergenic region upstream of ldhB, leads to activation of a second isozyme LDHB, which shows a strongly pH-dependent activity, overview
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additional information
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N-terminal deletion mutants lacking the first 5 and 10 amino acids of the N-terminus are more sensitive to denaturing environment than wild-type enzyme. They are easily inactivated and unfolded. Their instability increases and their ability to refold decreases with the increased number of amino acid residues removed from the N-terminus of LDH
additional information
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enhanced stability of L-lactate dehydrogenase through immobilization engineering, the enzyme is immobilized on glyoxyl-agarose, method optimization: preparation of an active and highly stable immobilizedderivative of LDH. with 90.1% immobilization and 72.0% yield is achieved using 300 mM trehalose during the immobilization process. Thermal stabilization factors attained for the immobilized LDH are 1600times greater as compared to its soluble counterpart. The immobilized preparation is also stabilized against ethanol where it recovers 75% of its initial activity after 48 h while the soluble enzyme is completely inactivated after only 10 min under the same conditions. Production of L-lactic acid is achieved in a batch reactor with the immobilized LDH and this preparation resists 15 reuses without the loss of activity. Co-immobilization of LDH and formate dehydrogenase. Immobilization has a negligible effect on the catalytic performance of L-LDH while having a significant impact on its stability
additional information
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