Team:UC Davis/Protein Engineering MM

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UC Davis iGEM 2014

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Experimentally Derived Kinetic Constants for Our Aldehyde Dehydrogenases

Now that we had identified three aldehyde dehydrogenases with unique specificity profiles, we proceeded to experimentally determine the kinetic constants (kcat and KM) of these enzymes on each substrate in our assay conditions.

Kinetic Constants of WT Escherichia coli Aldehyde Dehydrogenase
Substratekcat (s-1)kcat Error (s-1)KM (uM)KM Error (uM)Ki (uM)Ki Error (uM)kcat/KM (M-1s-1)kcat/KM Error (M-1s-1)
Propionaldehyde80.27782.8750.83N/AN/A10221.04748
Butyraldehyde6.270.28196.6827.65N/AN/A31867.744700.95
Pentanal6.180.2245.926.77N/AN/A13459820398.49
Hexanal9.0212.14335.75419.889754.199416.623252307.4341152614.3748
HeptanalN/AN/AN/AN/AN/AN/A645803940
Octanal4.530.6140.3323.41N/AN/A112265.5266927.82
Nonanal3.990.4427.6214.1N/AN/A144584.4275534.7
DecanalN/AN/AN/AN/AN/AN/A501203230
E-2-Pentenal0.940.0599.3320.74N/AN/A9480.512054.66
E-2-Hexenal1.290.0431.164.84N/AN/A41478.416599.45
E-2-Octenal0.990.0312.271.86N/AN/A81100.4712529.78
E-2-Nonenal0.870.029.61.16N/AN/A90621.9311127.45
E-2-Decenal0.660.0113.341.37N/AN/A49715.955217.05
E,E-2,4-Heptadienal0.680.11142.0875.99N/AN/A4809.312682.89
E,E-2,4-NonadienalN/AN/AN/AN/AN/AN/A1569221.9
E,E-2,4-DecadienalNDNDNDNDNDNDNDND
Kinetic Constants of W176V Escherichia coli Aldehyde Dehydrogenase
Substratekcat (s-1)kcat Error (s-1)KM (uM)KM Error (uM)Ki (uM)Ki Error (uM)kcat/KM (M-1s-1)kcat/KM Error (M-1s-1)
Propionaldehyde7.571.242575.71559.82N/AN/A2937.45800.31
Butyraldehyde4.420.12532.7032.00N/AN/A8306.37546.34
Pentanal5.440.35165.3535.25N/AN/A32877.937328.72
Hexanal7.290.3738.938.48N/AN/A187159.1441828.31
Heptanal12.461.76100.8923.73591.10152.70123500.8433883.89
Octanal16.215.97301.64146.69547.52335.1853752.8232794.34
Nonanal10.451.88161.8345.711292.60584.8964574.7321618.61
Decanal6.880.7191.5417.331764.75570.9975154.8816187.92
E-2-PentenalNDNDNDNDNDNDNDND
E-2-Hexenal0.220.0228.119.85N/AN/A7797.242796.55
E-2-OctenalN/AN/AN/AN/AN/AN/A2056.70237.60
E-2-NonenalN/AN/AN/AN/AN/AN/A1554.50261.50
E-2-DecenalN/AN/AN/AN/AN/AN/A1299.10210.50
E,E-2,4-HeptadienalNDNDNDNDNDNDNDND
E,E-2,4-NonadienalNDNDNDNDNDNDNDND
E,E-2,4-DecadienalNDNDNDNDNDNDNDND
Kinetic Constants of WT Rattus norvegicus (Rat) Aldehyde Dehydrogenase
Substratekcat (s-1)kcat Error (s-1)KM (uM)KM Error (uM)Ki (uM)Ki Error (uM)kcat/KM (M-1s-1)kcat/KM Error (M-1s-1)
Propionaldehyde19.90.511002.3645.02N/AN/A19856.171026.79
Butyraldehyde12.460.22261.0213.6N/AN/A47740.022629.46
Pentanal10.510.44118.0517.28N/AN/A89060.7913565.38
Hexanal8.280.6632.911.62N/AN/A251744.0991148.58
HeptanalN/AN/AN/AN/AN/AN/A859347269
Octanal6.620.57107.9432.67N/AN/A61315.1819282.67
Nonanal7.620.6888.6929.02N/AN/A85930.0629145.22
Decanal9.060.6693.8424.92N/AN/A96541.826598.58
E-2-Pentenal8.680.54694.0586.75N/AN/A12509.321745.41
E-2-Hexenal8.540.15220.1411.54N/AN/A38785.272139.18
E-2-Octenal7.510.4765.8116.08N/AN/A114129.5628807.7
E-2-Nonenal9.50.5875.4117.52N/AN/A125987.9430284.84
E-2-Decenal6.050.2558.699.63N/AN/A103066.4517434.71
E,E-2,4-Heptadienal6.010.3131.777.4N/AN/A189320.3945208.07
E,E-2,4-Nonadienal6.440.4749.3814.83N/AN/A130374.0340304.11
E,E-2,4-Decadienal6.320.3330.757.27N/AN/A205360.2549735.07

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