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Michaelis-Menten Now that we had identified four 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
Substrate	kcat	kcat Error	KM	KM Error	kcat/KM	kcat/KM Error
Propionaldehyde	8.00	0.27	782.87	50.83	10221.04	748.00
Butyraldehyde	6.27	0.28	196.68	27.65	31867.74	4700.95
Pentanal	6.18	0.22	45.92	6.77	134598.00	20398.49
Hexanal	5.37	0.53	12.65	6.36	424348.82	217516.13
Heptanal	Bad Fit	---	---	---	---	---
Octanal	4.53	0.61	40.33	23.41	112265.52	66927.82
Nonanal	3.99	0.44	27.62	14.10	144584.42	75534.70
Decanal	Bad Fit	---	---	---	---	---
E-2-Pentenal	0.94	0.05	99.33	20.74	9480.51	2054.66
E-2-Hexenal	1.29	0.04	31.16	4.84	41478.41	6599.45
E-2-Octenal	0.99	0.03	12.27	1.86	81100.47	12529.78
E-2-Nonenal	0.87	0.02	9.60	1.16	90621.93	11127.45
E-2-Decenal	0.66	0.01	13.34	1.37	49715.95	5217.05
E,E-2,4-Heptadienal	0.68	0.11	142.08	75.99	4809.31	2682.89
E,E-2,4-Nonadienal	Bad Fit	---	---	---	---	---
E,E-2,4-Decadienal	ND	ND	ND	ND	ND	ND

Kinetic Constants of W176V Escherichia coli Aldehyde Dehydrogenase
Substrate	kcat	kcat Error	KM	KM Error	kcat/KM	kcat/KM Error
Propionaldehyde	7.57	1.24	2575.71	559.82	2937.45	800.31
Butyraldehyde	4.42	0.12	532.70	32.00	8306.37	546.34
Pentanal	5.44	0.35	165.35	35.25	32877.93	7328.72
Hexanal	7.29	0.37	38.93	8.48	187159.14	41828.31
Heptanal	5.87	0.38	26.17	7.87	224363.85	68987.72
Octanal	6.55	0.45	84.25	21.41	77758.66	20464.40
Nonanal	6.41	0.31	75.29	13.54	85145.04	15837.70
Decanal	4.79	0.18	47.93	7.32	99954.31	15711.24
E-2-Pentenal	ND	ND	ND	ND	ND	ND
E-2-Hexenal	0.22	0.02	28.11	9.85	7797.24	2796.55
E-2-Octenal	Bad Fit	---	---	---	---	---
E-2-Nonenal	Bad Fit	---	---	---	---	---
E-2-Decenal	Bad Fit	---	---	---	---	---
E,E-2,4-Heptadienal	ND	ND	ND	ND	ND	ND
E,E-2,4-Nonadienal	ND	ND	ND	ND	ND	ND
E,E-2,4-Decadienal	ND	ND	ND	ND	ND	ND

Kinetic Constants of WT Rattus norvegicus (Rat) Aldehyde Dehydrogenase
Substrate	kcat	kcat Error	KM	KM Error	kcat/KM	kcat/KM Error
Propionaldehyde	19.90	0.51	1002.36	45.02	19856.17	1026.79
Butyraldehyde	12.46	0.22	261.02	13.60	47740.02	2629.46
Pentanal	10.51	0.44	118.05	17.28	89060.79	13565.38
Hexanal	8.28	0.66	32.90	11.62	251744.09	91148.58
Heptanal	Bad Fit	---	---	---	---	---
Octanal	6.62	0.57	107.94	32.67	61315.18	19282.67
Nonanal	7.62	0.68	88.69	29.02	85930.06	29145.22
Decanal	9.06	0.66	93.84	24.92	96541.80	26598.58
E-2-Pentenal	8.68	0.54	694.05	86.75	12509.32	1745.41
E-2-Hexenal	8.54	0.15	220.14	11.54	38785.27	2139.18
E-2-Octenal	7.51	0.47	65.81	16.08	114129.56	28807.70
E-2-Nonenal	9.50	0.58	75.41	17.52	125987.94	30284.84
E-2-Decenal	6.05	0.25	58.69	9.63	103066.45	17434.71
E,E-2,4-Heptadienal	6.01	0.31	31.77	7.40	189320.39	45208.07
E,E-2,4-Nonadienal	6.44	0.47	49.38	14.83	130374.03	40304.11
E,E-2,4-Decadienal	6.32	0.33	30.75	7.27	205360.25	49735.07