On line supplementary material
The pH dependence of kinetic isotope effects in monoamine oxidase A indicates
stabilisation of the neutral amine in the enzyme-substrate complex.
Rachel V. Dunn, Ker Marshall, Andrew W. Munro, Nigel S. Scrutton
Faculty of Life Sciences, Manchester Interdisciplinary Biocentre, University of
Manchester, 131 Princess Street, Manchester, M1 7DN, UK.
0.16
0.14
A
0.12
-1
kcat (s )
0.10
0.08
0.06
0.04
0.02
0.00
6.5
7.0
7.5
8.0
8.5
9.0
9.5
pH
1.8
B
1.6
-1
-1
kcat/Km (s mM )
1.4
1.2
1.0
0.8
0.6
0.4
0.2
0.0
7.0
7.5
8.0
8.5
9.0
9.5
pH
Fig S1. pH dependence of the steady-state kinetic parameters of MAO A-catalysed
oxidation of benzylamine at 20 oC. (A) Plot of kcat versus pH. (B) Plot of kcat/Km
versus pH.
0.10
0.09
A
0.08
0.07
-1
kred (s )
0.06
0.05
0.04
0.03
0.02
0.01
0.00
6.5
7.0
7.5
8.0
8.5
9.0
9.5
8.5
9.0
9.5
pH
3.0
B
2.0
-1
kred/Ks (s mM )
2.5
-1
1.5
1.0
0.5
0.0
6.5
7.0
7.5
8.0
pH
Fig. S2. pH dependence of the reductive half-reaction of MAO A-catalysed oxidation
of benzylamine at 20 oC. (A) Plot of kred versus pH. (B) Plot of kred/Ks versus pH.
0.09
0.085
Relative absorbance
0.08
0.075
0.07
0.065
0.06
0.055
0.05
0.045
0.04
0
2
4
6
8
10
12
14
16
18
20
Time (s)
Fig. S3. Reaction transient for MAO A-catalysed oxidation of 0.5 mM PEA at pH
9.0, 20 oC. The data (black line) were fit to an equation for a double exponential
decay with offset (grey line).
0.07
0.065
Relative absorbance
0.06
0.055
0.05
0.045
0.04
0.035
0.03
0.025
0.02
0
20
40
60
80
100
120
140
160
180
200
Time (s)
Fig. S4. Reaction transient for MAO A-catalysed oxidation of 0.4 mM benzylamine
at pH 8.5, 20 oC.
The data (black line) were fit to an equation for a double
exponential decay with offset (grey line).
3.5
3.0
2.5
-1
k (s )
2.0
1.5
1.0
0.5
0.0
0.0
0.5
1.0
1.5
2.0
PEA (mM)
Fig. S5. Substrate dependence of the reductive half-reaction of MAO A-catalysed
oxidation of PEA at pH 8.5, 20 oC. The rate at each concentration is the average of at
least three replicates, and the error is the standard deviation. The data were fit to the
equation y = (kcat*x)/(Ks+x); the errors in the limiting rate of flavin reduction and the
substrate binding constant are those determined from curve fitting.