1.1.1.67: mannitol 2-dehydrogenase
This is an abbreviated version!
For detailed information about mannitol 2-dehydrogenase, go to the full flat file.
Word Map on EC 1.1.1.67
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1.1.1.67
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polyol
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fluorescens
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d-fructose
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synthesis
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5-dehydrogenase
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1.1.1.138
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heterofermentative
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reuteri
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gluconobacter
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industry
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pfldh
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arabitol
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molasses
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nutrition
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biotechnology
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analysis
- 1.1.1.67
-
polyol
- fluorescens
- d-fructose
- synthesis
-
5-dehydrogenase
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1.1.1.138
-
heterofermentative
- reuteri
- gluconobacter
- industry
- pfldh
- arabitol
- molasses
- nutrition
- biotechnology
- analysis
Reaction
Synonyms
alcohol dehydrogenase, zinc-containing, D-mannitol dehydrogenase, M2DH, mannitol 2-dehydrogenase, mannitol dehydrogenase, mannitol-2-dehydrogenase, MDH, mt-dh, MtDH, MtlD, NAD+-dependent mannitol dehydrogenase, NADH-dependent mannitol dehydrogenase, pfMDH, polyol dehydrogenase, PsM2DH, TM0298, TM_0298
ECTree
Advanced search results
Engineering
Engineering on EC 1.1.1.67 - mannitol 2-dehydrogenase
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D230A
the mutant shows reduced catalytic efficiency compared to the wild type enzyme
D69A
E133A
the mutant shows reduced catalytic efficiency compared to the wild type enzyme
E133Q
the mutant shows reduced catalytic efficiency compared to the wild type enzyme
E292A
mutation partially disrupts the catalytic cycle. Role for residue Glu292 as a gate in a water chain mechanism of proton translocation. Removal of gatekeeper control in the E292A mutant results in a selective, up to 120fold slowing down of microscopicsteps immediately preceding catalytic oxidation of mannitol, consistent with the notion that formation of the productive enzyme-NAD-mannitol complex is promoted by a corresponding position change of Glu292
E68K
site-directed mutagenesis, the mutant shows an altered cofactor specificity compared to the wild-type enzyme, which is switched to NADP(H), EC 1.1.1.138, NADP(H) is preferred by 10fold over NAD(H)
E68K/D69A
shows about a 10fold preference for NADP(H) over NAD(H), accompanied by a small decrease in catalytic efficiency for NAD(H)-dependent reactions as compared to wild-type enzyme
H303A
H303A/R373A/K381A
the mutant shows severely reduced catalytic efficiency compared to the wild type enzyme
K295A
K295M
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2000000fold lower turnover number for D-mannitol oxidation at pH 10.0 than the wild-type enzyme
K381A
the mutant shows severely reduced catalytic efficiency compared to the wild type enzyme
N191A
N191A/N300A
the rate constants for the overall hydride transfer to and from C-2 of mannitol are selectively slowed, with additive effects in the double mutant
N191D
the internal equilibrium of enzyme-NADH-fructose and enzyme-NAD+-mannitol is altered 10000- to 100000fold from being balanced in the wild-type enzyme to favoring enzyme-NAD+-mannitol in the single site mutants, N191D and N300D. N191D and N300D appear to lose fructose binding affinity due to deprotonation of the respective Asp above apparent pK values of 5.3 0.1 and 6.3 0.2, respectively
N191D/N300D
mutant behaves as a slow fructose reductase at pH 5.2, lacking measurable activity for mannitol oxidation in the pH range 6.8-10
N191L
the rate constants for the overall hydride transfer to and from C-2 of mannitol are selectively slowed, between 540- and 2700fold. Partial disruption of the oxyanion hole in the single-site mutant causes an upshift, by about 1.2 pH units, in the kinetic pK of the catalytic acid-base Lys295 in the enzymeNAD+-mannitol complex
N300A
N300D
N300S
the mutant shows severely reduced catalytic efficiency compared to the wild type enzyme
R373A
the mutant shows severely reduced catalytic efficiency compared to the wild type enzyme
additional information
site-directed mutagenesis, the mutant shows an altered cofactor specificity compared to the wild-type enzyme, which is switched to NADP(H), EC 1.1.1.138, NADP(H) is equally utilized as NAD(H)
D69A
utilizes NAD(H) and NADP(H) with similar catalytic efficiencies. Uses NADP(H) almost as well as wild-type enzyme uses NAD(H)
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mutant enzyme displays catalytic efficiency for NAD+-dependent oxidation of D-mannitol 300fold below the wild-type value
H303A
the mutant shows severely reduced catalytic efficiency compared to the wild type enzyme
K295A
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30000fold lower turnover number for D-mannitol oxidation at pH 10.0 than the wild-type enzyme
K295A
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mutant enzyme displays catalytic efficiency for NAD+-dependent oxidation of D-mannitol 400000fold below the wild-type value
the rate constants for the overall hydride transfer to and from C-2 of mannitol are selectively slowed, between 540- and 2700fold. Partial disruption of the oxyanion hole in the single-site mutant causes an upshift, by about 1.2 pH units, in the kinetic pK of the catalytic acid-base Lys295 in the enzymeNAD+-mannitol complex
N191A
the mutant shows severely reduced catalytic efficiency compared to the wild type enzyme
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mutant enzyme displays catalytic efficiency for NAD+-dependent oxidation of D-mannitol 1000fold below the wild-type value
N300A
the mutant shows severely reduced catalytic efficiency compared to the wild type enzyme
the internal equilibrium of enzyme-NADH-fructose and enzyme-NAD+-mannitol is altered 10000- to 100000fold from being balanced in the wild-type enzyme to favoring enzyme-NAD+-mannitol in the single site mutants, N191D and N300D. N191D and N300D appear to lose fructose binding affinity due to deprotonation of the respective Asp above apparent pK values of 5.3 0.1 and 6.3 0.2, respectively
N300D
the mutant shows severely reduced catalytic efficiency compared to the wild type enzyme
D-mannitol production by resting state whole cell biotransformation of D-fructose by heterologous mannitol dehydrogenase gene from Leuconostoc pseudomesenteroides and the formate dehydrogenase gene, gene fdh from Mycobacterium vaccae N10, expression in Bacillus megaterium, development of an in vivo system, overview
additional information
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D-mannitol production by resting state whole cell biotransformation of D-fructose by heterologous mannitol dehydrogenase gene from Leuconostoc pseudomesenteroides and the formate dehydrogenase gene, gene fdh from Mycobacterium vaccae N10, expression in Bacillus megaterium, development of an in vivo system, overview
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additional information
application of a modular screening procedure that can identify the optimal operating policy of an enzymatic reactor, which minimizes the enzyme consumption, given the process kinetic model, and an imposed production capacity. Following an optimization procedure, the process effectiveness is evaluated in a systematic approach, by including simple batch reactor (BR), batch with intermittent addition of the key-enzyme following certain optimal policies (BRP). The enzymatic reduction of D-fructose to mannitol is used as a model system utilizing suspended MDH (mannitol dehydrogenase) and NADH (nicotinamide adenine dinucleotide) cofactor, with the in-situ continuous regeneration of the cofactor by the expense of formate degradation in the presence of suspended FDH (formate dehydrogenase). The NADH-dependent FDH and MDH typical activity in D-fructose reduction is of 1-2 U/ml in a batch reactor
additional information
Pseudomonas fluorescens DSM 50106
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application of a modular screening procedure that can identify the optimal operating policy of an enzymatic reactor, which minimizes the enzyme consumption, given the process kinetic model, and an imposed production capacity. Following an optimization procedure, the process effectiveness is evaluated in a systematic approach, by including simple batch reactor (BR), batch with intermittent addition of the key-enzyme following certain optimal policies (BRP). The enzymatic reduction of D-fructose to mannitol is used as a model system utilizing suspended MDH (mannitol dehydrogenase) and NADH (nicotinamide adenine dinucleotide) cofactor, with the in-situ continuous regeneration of the cofactor by the expense of formate degradation in the presence of suspended FDH (formate dehydrogenase). The NADH-dependent FDH and MDH typical activity in D-fructose reduction is of 1-2 U/ml in a batch reactor
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