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Article title: Ligninase-mediated Transformation of 4, 4’-Dibromodiphenyl Ether
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Journal name: Environmental Science and Pollution Research
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Authors: Yiping Feng \and Liang Mao \and Yijun Chen \and Shixiang Gao
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Address: Yiping Feng \and Liang Mao \and Yijun Chen \and Shixiang Gao \ at State Key
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Laboratory of Pollution Control and Resource Reuse, School of the Environment, Nanjing
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University, Nanjing 210023, PR China
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The e-mail address, telephone and fax numbers of the corresponding author (Shixiang Gao):
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\ Tel.: +86 -25 -89680359, Fax: +86 -25 -89680359 \\ E-mail: ecsxg@nju.edu.cn
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Supplemental Material:
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Content:
Schematic depiction of LiP catalytic cycle (Fig.
SM-1
Introduction
SM-1)
The discussion of Enzymatic reaction kinetics (Fig.
SM-2
Results and Discussion
SM-2)
Effect of VA and NOM (Fig. SM-3) and Table SM-1
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1
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Introduction
Fig. SM-1:
VA
Native LiP
LiP-Ⅲ
(6)
H2O2
RH
(4)
(1)
(3)
H2O
RH2
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6
7
8
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Permanent Deactivation
LiP-Ⅱ
RH2
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Excess H2O2
(2)
LiP-Ⅰ
(5)
RH
Fig. SM-1 Schematic depiction of LiP catalytic cycle. The steps 1, 2 and 3 (in red) represent the
catalytic cycle and the steps 4 and 5 (in black) denote LiP inactivation via LiPIII and step 6 (in
blue) represent the mitigation of LiP inactivation in the presence of VA. (adapted from Mao et
al.2010)
Results and Discussion
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Enzymatic reaction kinetics
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The Michaelis-Menten equation shows the relationship between initial velocity of
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enzymatic reaction and substrate concentration (Fig. SM-2.A). The simplified form can be
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described as:
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V
V max [ S ]
,
m  [ S ]
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where V is the initial reaction rate for each initial substrate concentration and Km is the substrate’s
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Michaelis constant, Vmax corresponds to the maximum reaction rate which can be achieved when
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substrate concentration is sufficiently to saturate all available enzymes. Usually, the conversion
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form of Michaelis-Menten equation, the Lineweaver-Burk equation:
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1
m 1
1
,


V V max [ S ] V max
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is applied to derive K m and Vmax. 1/V plot of 1/[S], obtained a straight line (Fig. SM-2.B).
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The absolute value of straight line intercept with the x-axis was 1/K m ; and 1/Vmax was
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straight line intercept with the y-axis.
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Fig. SM-2:
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(A)
0.45
0.4
0.35
Rate, nM/s
0.3
0.25
0.2
0.15
0.1
0.05
0
0
100
200
300
400
500
BDE15 Concentration, nM
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(B)
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y = 1515.x + 2.114
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1/v
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y = 497.5x + 1.739
4
2
0
-0.01
-0.005
-2 0
-4
0.005
0.01
1/s
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4
0.015
0.02
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Fig. SM-2. Michaelis-Menten curves (A) and Lineweaver-Burk equation (B) for LiP-catalyzed
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BDE 15 reactionsExperiment conditions: [BDE 15] = 381.1 nM; [LiP] = 0.047 U mL-1, [H2O2] =
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0.2 mM, [VA] = 2 mM, pH = 4 (10-mM CPBS), Reaction time = 40 seconds. Error bars represent
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standard deviations (n=3). Without NOM (in balck), with 8.41 mg (TOC) L-1 NOM (in red).
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Effect of VA and NOM on LiP-mediated BDE 15 transformation
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Fig. SM-3:
0.25
[NOM] 0mg/L
[NOM] 4.14mg/L
[NOM] 8.41mg/L
[NOM] 12.68mg/L
Absorbance
0.2
0.15
0.1
0.05
0
0
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300
400
500
600
Time（seconds）
Fig. SM-3. Rate of VA conversion by LiP in reaction system containing NOM at various
concentration.
Experiment conditions: [NOM] = 0, 4.14, 8.41 and 12.68 mg (TOC) L-1. [LiP] = 0.031 U mL-1,
[H2O2] = 0.2 mM, pH = 4 (10-mM CPBS), [VA] = 2 mM.
Table SM-1:
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Kinetics parameter of BDE 15 transformation in LiP-mediated reaction system that containing VA
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and/or NOM. Experiment conditions: [LiP] = 0.047 U mL-1, [H2O2] = 0.2 mM, pH = 4 (10-mM
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CPBS), [VA] = 2 mM, [NOM] = 8.41 mg (TOC) L-1. Error bars represent standard deviations
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(n=3).
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Treatment groups
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a
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b
k0 (min-1)a
y=k0x+b
R2
without VA or NOM
-2.02±0.27b
Y=-2.02x+341.26
0.97
with NOM
-1.99±0.16b
Y=-1.99x+342.22
0.97
with VA
-5.45±0.28
Y=-5.45x+340.28
0.99
with VA and NOM
-4.19±0.05
Y=-4.19x+346.83
0.98
Pseudo-first-order kinetic rate constant for LiP-mediated transformation of BDE 15.
No difference value was significant between the two (p < 0.05), and for others, differences
between treatment groups were considered statistically significant at p < 0.05.
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