James Lee Enzyme Kinetics Solution PDF

BK10110302 V.
PRASARNTH RAAJ VEERA RAO – BIOPROCESS
Solution 2.1
Total volume = 44 + 5 + 1= 50ml
From the graph the equation obtained
y=0.033× + 0.03a 1
m= 0.033 ɱ mol / ml . min
a) Activity of the β- glucosidase
0.033 x 50= 1.65 mumol / min
i) = 1.65mumol/min
0.1 mg/ml x 0.1ml
= 165 units/mg protein
ii) = 1.65 mumol/min

1ml of enzyme
= 1.65 units/ml of enzyme
b) Initial rate of reaction 0.033 mumol/ mL.min
S vs t Graph
1.2
1 y = 0.033x + 0.0391
0.8
0.6
0.4
0.2
0
0 5 10 15 20 25 30 35
BK10110302 V.PRASARNTH RAAJ VEERA RAO – BIOPROCESS
Solution 2.2
→
←
→
←
a) Michaelis-Menten approach
The rate of product formation.
d[p]
( )
dt
Since the enzyme is preferred,
Make E as the subject,
Since forward reaction = backward reaction.
( )
ubstitute into :
[( ) ( ) ( )] ( )
( ) ( )( ) ( )( ) ( )( )
Make as a subject:
( ) ( )( ) ( )( )
( )( ) ( )( )
( )
( ) [( )( )]
( )( )
( )
( ) ( )( )
ub into :
( )( ) ( )( )
( )
( )( )( ) ( )( )( )
( )
Make as a subject,
( )( )( )
( )( ( )( ))
( )( )( )
( )
( )( ( )( ))
( )( )( )
( )
( )
( ) ( )( )
ub into ,
d[p] ( )( )( )
( )
dt ( )
( )( )
( )( )( )
)
( )
( )( )
Since ( )
( )( )
)
( )
( )( )
b) Since [ ] [ ]
d[p] ( )( )
dt ( )( )
Solution 2.3
(a) E+S k1 (ES)1
(ES)1 k3 (ES)2
(ES)2 k2 E+P
[ ]
V= = k5 [ES] 2
[E0] = [E] + [ES] + [ES]2
[E] = [E0]-[ES]-[ES]2
k2 = [E] [S]
k1 [ES]1
k2 [ES]1 = [Eo] [S] – [ES]1 [S]

k1
[ES]1 ( k2 + [S] ) = [E0] [S] – [ES]2 [S]

k1
[ES]1 = [E0] [S] – [ES]2 [S]
k2 + [S]
k1
k4 = [ES]1
k3 [ES]2
k4 [ES]2 = [E0] [S] – [ES]2 [S]

k3 k2 + [S]
k1
[ES]2 ( k2 k4 + k4 [S] ) = [E0] [S] – [ES]2 [S]
k1k3 k3
[ES]2 ( k2 k4 + k4 [S] + [S] )= [E0] [S]

k1 k3 k3
[ES]2 = [E0] [S]

k2 k4 + k4 [S] + [S]
k1 k3 k3
V= d [P] = k5 [E0] [S]

dt k2 k4 + k4 [S] + [S]
k1 k3 k3
= Vm [S]
k2 k4 + k4 [S] + [S]
k1 k3 k3
Solution 2.4
a) Michaelis-Menten approach
The rate of product formation.
d[p]
( )
dt
Make E as the subject,
Since forward reaction = backward reaction.
( )
ubstitute into :
[( ) ( ) ( )] ( )
( ) ( )( ) ( )( ) ( )( )
Make as a subject:
( ) ( )( ) ( )( )
( )( ) ( )( )
( )
( ) [( )( )]
( )( )
( )
( ) ( )( )
ub into :
( )( ) ( )( )
( )
( )( )( ) ( )( )( )
( )
Make as a subject,
( )( )( )
( )( ( )( ))
( )( )( )
( )
( )( ( )( ))
( )( )( )
( )
( )
( ) ( )( )
ub into ,
d[p] ( )( )( )
( )
dt ( )
( )( )
( )( )( )
)
( )
( )( )
Since ( )
( )( )
)
( )
( )( )
b) Briggs-Haldane approach
→
←
→
←
→
←
The rate of product formation,
d(p)
( )
dt
Make as a subject,
Substrate consumption,
d( )
( )( ) ( )( ) ( ) ( )
dt
d( )
( )( ) ( ) ( )
dt
ubstitute into :
( )( ) ( ) ( )( ) ( )
( )( ( ) ) ( )( ) ( )
( )( ( ) ) ( ) ( )( ) ( )( ) ( )
( )( ( ) ( )) ( ) ( )( ) ( )
( ) ( )( ) ( )
( )
( ( ) ( ))
ubstitute into
( )( ) ( )( )
( ) ( )( ) ( )
( )( ) )( )
( ( ) ( ))
( ) ( ) ( ) ( )( ) ( ) ( )
( )( ) )
( ( ) ( ))
( )( )( ( ) ( ))
( ) ( ) ( ) ( )( ) ( ) ( )
( )( ( ) ( ) ( ) )( ( ) ( )) ( ) ( )
( ) ( )
( )
( ( ) ( ) ( ) )( ( ) ( ))
ubstitute into
d(p) ( ) ( )
(
dt ( ( ) ( ) ( ) )( ( ) ( ))
d(p) v ( ) ( )
dt ( ( ) ( ) ( ) )( ( ) ( ))
Solution 2.5
Lineweaver- Burk Plot
x-intercept= - 1
km
y-intercept= 1/ V more
Equation obtained y= 0.0172 x + 3.6342
y-intercept = 3.6342= 1/ V max
V max = 0.275
x-intercept , y= 0
0.0172x + 3.6342=0
0.0172x = -3.6342
x= -211.291
x= -1/km
km = 1/211.291
= 0.00473
Longmuir Plot
Equation obtained y= 3.3133x + 0.0191
1/Vm = m = 3.3133
Vm=0.302
y-intercept= km/Vm = 0.0191
Km = 0,0191x 0.302
= 0.00577
Eadie-Hofstee Plot
Equation obtained y= -0.0043x + 0.2645
-Km = m = -3.3133
Km=0.302
y-intercept= Vm = 0.2645
Non-Linear Regression Procedure
From the graph, Vm=0.2
½ Vmax = 0.1, Km=0.0032
Data for Graph plot :
Langmuir Plot
s s/v
0.0032 0.028829
0.0049 0.033108
0.0062 0.043357
0.008 0.048193
0.0095 0.0475
Lineweaver-Burk Plot
1/s 1/v
312.5 9.009009
204.0816 6.756757
161.2903 6.993007
125 6.024096
105.2632 5 Kinetic Parameters
Type of Plot
Vmax Km
Eadie-Hofstee Plot
Langmuir 0.2750 0.0047
v/s v Lineweaver-Burk 0.0191 0.0057
34.6875 0.111 Eadie-Hofstee 0.2645 0.0043
30.20408 0.148 Non-Linear Regression 0.2000 0.0032
23.06452 0.143
20.75 0.166
21.05263 0.2
Non-Linear Regression Plot

S v
0.0032 0.111
0.0049 0.148
0.0062 0.143
0.008 0.166
0.0095 0.2
Langmuir Plot
0.06
0.05 y = 3.3133x + 0.0191
0.04
0.03
0.02
0.01
0
0 0.002 0.004 0.006 0.008 0.01
Lineweaver Burk Plot

10
9 y = 0.0172x + 3.6342
8
7
6
5
4
3
2
1
0
0 50 100 150 200 250 300 350
Eadie-Hofstee Plot
0.25
0.2
0.15
y = -0.0043x + 0.2645
0.1
0.05
0
0 5 10 15 20 25 30 35 40
Non-Linear Regression Procedure

0.25
0.2
0.15
0.1
0.05
0
0 0.002 0.004 0.006 0.008 0.01
Solution 2.6
Rate of product formation

( )
v ( )
Enzyme is preserved,
( )
Assumptions:
( )
𝑑(𝐸𝑆)
d( ) [𝐸𝑂 ]small, 𝑑𝑡
neg igib e
dt
( )
( )( ) ( ) ( )
( )( ) ( )( )
( )( )
( ) ( )
Substitute equation into
( ( ))( )
( )
( )
( ) ( )
( )
( )
( )( ( ) ) ( )
( )
( )
( ( ) )
Substitute into
𝑣𝑚 𝑘 𝑘 𝐸𝑜
d(p) ( )
v ( )
dt ( ( ) ) 𝑘 𝑘
𝐾𝑚
( )
𝑘
( ( ) )
Dividing with the value of

( )
[( ( ) )
]/
v s
v
( ( ) )
Solution 2.7
a) FCs0 - FCs + rSv = V
For Batch reactor F=0
rSv = V
[ ]
=
[ ]
[ ]
=
[ ]
= 60mol/m3.min
b) Equation obtained y = 6.3852x + 59.571
m = Vmax = 6.3852
y- intercept = - Km = 59.571
Km = - 59.571
c) FCs0 - FCs + rSv = 0

FCs0 - FCs = - rSv = rpv
FCs0 - FCs = V
F = 0.0001m3/min
V = 0.0003m3
( FCSo - FCs ) (Km + Cs) = Vmax CsV

FCSo Km + FCSo Cs - FKm Cs – FCs = Vmax CsV
2
(0.0001 (300)(200) + 0.0001(300)Cs – 0.001(200)Cs – 0.001Cs2 = 100 (0.0003)Cs )

6 + 0.03Cs – 0.02Cs – 0.001Cs2 = 0.03Cs
0.0001Cs2 + 0.02Cs – 6 = 0
Cs=165mol/m3
Data :
Cs t t/ln(Cso/Cs) (Cso-Cs)/ln(Cso/Cs)
1 1 0.175322 52.42135
5 5 1.221197 72.0506
10 10 2.940141 85.26409
20 20 7.385387 103.3954
Graph :
(Cso-Cs)/ln(Cso/Cs)
120
y = 6.3852x + 59.571
100
80
(Cso-Cs)/ln(Cso/Cs)
60
Linear ((Cso-
40 Cs)/ln(Cso/Cs))
20
0
0 2 4 6 8
Solution 2.8
a) Km =0.01 mol/L
Cso = 3.4 x 10 -4 mol/L
Cs = 0.9 x 3.4 x 10-4
= 3.06 x 10-4 mol/L
t= 5minutes
[ ]
=
[ ]
( 3.4x 10-4 – 3.06 x 10-4) = Vmax (3.06 x 10-4)

S 0.01 + (3.06 x 10-4)
6.8 x 10-6 = Vmax ( 0.03)

Vmax = 2.27 x 10-4 mol/L-min
b) 6.8 x 10-6 x 15 = 1.02 x 10-4 mol/L

Solution 2.9
Km = 0.03mol/L
rmax = 13mol/L min × 60 = 780mol/L hr
F=10L/Hr
Cs=10mol/L
F=10L/Hr
Cs=0.5mol/L
CSTR
a) V = ?
CSTR @ Stead State

FCs0 - FCs + rSv = 0
F (Cs0 - Cs ) = rpv
( )
10 (10 – 0.5) = V
V = 0.129 liter
b) Plug - Flow @ Stead State
Km ln + (Cs0 - Cs ) = rmax t
0.03 ln + (10 - 0.5 ) = 780t

9.95899 = 780t
t = 0.0123hr
t = V/F = 0.0123
V = 0.0123 × 10
= 0.123liter
Solution 2.10
Km = 10g/L
rmax = 7g/L.min
a)
F=0.5L/min
Cs0=50g/L F=0.5L/min
1L Cs1=?g/L 1L
F=0.5L/min
Cs2=? g/L
CSTR @ Steady State

F (Cs0 - Cs ) = rpv
( s )
0.5 (50 – Cs1) = (1)
s
(25-0.5Cs1)(10+ Cs1)=7Cs1
250+25Cs1-5Cs1-0.5Cs12=7Cs1
0.5Cs12-13Cs1-250=0
Cs1=38.86g/L
( s )
0.5 (38.86 – Cs2) = (1)
s
(19.43-0.5 Cs2)(10+ Cs2)=7 Cs2
194.3+19.43Cs2-5Cs2-0.5Cs22=7Cs2
0.5Cs22-7.43Cs2-194.3 =0
Cs2=28.49g/L
b)
F=0.5L/min
Cs0=50g/L
2L
F=0.5L/min
Cs1=?g/L
CSTR @ Steady State

F (Cs0 - Cs ) = rpv
( s )
0.5 (50 – Cs1) = (2)
s
(25-0.5Cs1)(10+ Cs1)=14Cs1
250+25Cs1-5Cs1-0.5Cs12=14Cs1
0.5Cs12-6Cs1-250=0
Cs1=29.15g/L
Since in the Cs in two reactor system is less than Cs in one reactor system, therefore two reactor
system is more efficient than one reactor system as it indicates more substrates have been
consumed to form products.
Solution 2.11
a) k1 [E] [S] = k2 [ES]
[ES] = k2 k1 [E] [S]

k1 [E] [S] k2
k3 [E] [P] = k4 [EP]
[EP] = k3
k4 [E] [P]
k5 [ES][P] = k6 [ESP]
[ESP] = k5 [ES] [P]

k6
k7 [EP] [S] = k8 [ EPS ]

[EPS] = k7 [EP] [ S ]
K8
= k7 k3 [ S ]
k8 k4 [E] [P]
From,
[ESP] = k5 [P] k2
k6k1 [E] [S]
[E0] = [E] + [ES] + [EP] + [ESP] + [EPS]
[E0] = [ES] + [ESP] + [E] + [EP] + [EPS]

[ ][ ]
[E0] = [ES] + [ESP] + [E] + [EP] +
[ ]
[E0] = [ES] + [ESP] + [E] + [EP] + ( )
[ ][ ] [ ]
[E0] = [ES] + [ESP] + [E] + +( )
[ ] [ ]
[E0] = [ES] + [ESP] + [E] [ ( )]
[ ][ ] [ ][ ] [ ] [ ]
[E0] = [ES] + + [ ( )]
[ ] [ ] [ ] [ ]
[E0] = [ES] {1 + + [ ( )]}
[ES] = [ ] [ ] [ ] [ ]
[ ( )]
[ ]
V= = [ ] [ ] [ ] [ ]
[ ( )]
[ ][ ] [ ][ ]
b) KSP = KPS =
[ ] [ ]
[ ][ ][ ] [ ][ ][ ]
KSP = [ ]
KPS = [ ]
[ ][ ][ ] [ ][ ][ ]
[ESP] = [EPS] =
Given: [ESP] = [EPS]
KS KSP = KP KPS
[ ] [ ]
= [ ] [ ] [ ] [ ]
[ ]
[ ]
= [ ] [ ] [ ]
[ ] [ ]
c)
Ks=Kps 𝑘𝑠 𝑘𝑝𝑠 [ESP]=[EPS]
Kp=Ksp 𝑘𝑝 𝑘𝑠𝑝
[ ] ( )[ ]
( ) [ ]
[ ] [ ]
[ ][ ]
[ ] [ ][ ] [ ]
[ ][ ]
[ ]( [ ]) [ ]
[ ]
[ ]
( [ ])
[ ]
[ ]
( [ ])
Compare with
[ ]
[ ]
[ ]
Hence, Vmax = ( [ ])
[ ]
Km= [ ])
(
[ ] [ ]
d)
[ ]
*∫ [ ] ∫ +
[ ]
[ ]
* [ ] +
[ [ ] [ ]]
[ ] [ ] [ ] [ ]
[ ] [ ] [ ] [ ]
[ ]
[ ] [ ] n( )
[ ]
[ ] [ ]
n([ ] [ ])
[ ] [ ] [ ]
n( ) ( )
[ ]
Y = mx+c
[ ]
Y= n ([ ]
)
M=
[ ] [ ]
X= ( )
C=
[ ] [ ] [ ]
So we can plot a graph of n ([ ]
) vs ( )
Solution 2.14
Rate:
rp = k9CES +k10CEIS1 +k10CEIS2 ---- 1
Enzyme balance:
CEo = CE + CES ---- 2
CEo = CEIS1 + CES + CE ---- 3
CEo = CEIS2 + CEI + CE ---- 4
The equilibrium reaction equations are as follows:
CE Cs / CES = k2/k1 ---- 5 CECI / CEI = K4/K3 ---- 7
CESCI /CEIS1 = K6/K5 ---- 6 CEICS / CEIS2 = K8/K7 ---- 8
By rearranging Equation 5,
CE = (k2/k1) Cs CES
From Equation 2,
CEo = [(k2/k1)CE + 1] CES
 CES = CEo /[( k2/k1)CS +1] ---- 9
CES = [(K6/K5)CI ] CEIS1
From Equation 3,
CEo = CEIS +CES + (k2/k1) Cs CES
= {CEIS1 + [1 + (k2/k1) Cs]( K6/K5)CI }CEIS1
= {1 + [1 + (K2/K1) Cs ]( K6/K5)CI } CEIS1
 CEIS1 = CEo/ {1 + [1 + (k2/k1) Cs ]( K6/K5)CI } ---- 10
CE = (K4/K3) CEI
CEI = K8/K7CS CEIS2

From Equation 4,
CEo = CEIS2 + CEI + [(K4/K3)CI]CEI
= CEIS2 + [1 + (K4/K3)CI ]CEI
= CEIS2 + [1 + (K4/K3)CI ]( K8/K7)CS CEIS2
CEo = {1 + [1 + (K4/K3)CI ]( K8/K7)CS } CEIS2
 CEIS2 = CEo / {1 + [1 + (K4/K3)CI ]( K8/K7)CS } ---- 11
From Equation 1, since rp = k9CES +k10CEIS1 +k10CEIS2,
By substituting Equation 9, 10 & 11 into Equation 1,
Therefore,
rp = k9 CEo /[( k2/k1)CS +1] + k10 CEo/ {1 + [1 + (k2/k1) Cs ]( K6/K5)CI } + k10 CEo / {1 + [1 + (K4/K3)CI
]( K8/K7)CS }
Solution 2.15
a) Based on the graphs

The y-intercept in Lineweaver – Burk plot is almost the same.
Y-intercept => 3.8266; 3.6342

Whereas in Langmuir Plot
Two equations obtained
Y = 2.9883x + 0.0489
Y = 3.3133x + 0.0191
When y=0
X= ; X=
X = -0.016 ; X = -0.005
In Line weaver – Burk Plot and Langmuir Plot both indicates it’s a competitive inhibitor
Data :
Lineweaver
1/s 1/Vo 1/Vi
312.5 9.009009 16.94915
204.0816 6.756757 14.08451
161.2903 6.993007 10.98901
125 6.024096 9.009009
105.2632 5 8
Langmuir
s s/Vo S/Vi
0.0032 0.028829 0.054237
0.0049 0.033108 0.069014
0.0062 0.043357 0.068132
0.008 0.048193 0.072072
0.0095 0.0475 0.076
Lineweaver-Burk Plot
20
18 y = 0.0439x + 3.8266
16
14
1/Vo
12
10 1/Vi
y = 0.0172x + 3.6342
8 Linear (1/Vo)
6 Linear (1/Vi)
4
2
0
0 100 200 300 400
Langmuir Plot
0.09
0.08
y = 2.9883x + 0.0489
0.07
0.06 s/Vo
0.05 y = 3.3133x + 0.0191
S/Vi
0.04
Linear (s/Vo)
0.03
Linear (S/Vi)
0.02
0.01
0
0 0.002 0.004 0.006 0.008 0.01
b) Y-intercept = 1/Vmax = 0.00489

Vmax = 1/0.00489 = 204.5 mol /L.min
Km/Vmax = 2.9883
Km=2.9883*204.5
=611mol/L
Solution 2.16
(a) E + S ↔
ES + S ↔
→k5 E + P
(E0 ) = (E) +(ES) + (ESS)

(E) = (E0 ) – (ES) – (ESS) --------
( )
V= = k5 (ES) ------
( )( )
= ( )
( ) = (ES)(S)/ (k4 / k3)
K2 / k1 = (E)(S) / (ES)
K2/k1 (ES) = (E0)(S) – (ES)(S)
(ES)((k2/k1) + (S)) = (E0)(S) – (ES)(S)2 /
(ES)( (k2/k1) + (S)( ) ) = (E0)(S) – (ES)(S)2
(ES) ( (k2/k1) + (S)( ) + (S)2 ) = (E0)(S)
(ES) = (E0)(S) / (k2/k1) + (S)( ) + (S)2 --------
3→
( )
V= = k5 (E0) (S) / (k2/k1) + (S)( ) + (S)2
= Vm(S) / (k2/k1) + (S)( ) + (S)2
(b) At low substrate concentration,

1/ Vm = 3.1209
Vm = 0.3204
Km/Vm = 106.07
Km / 0.3204 = 106.7
K1m = 33.98
At high substrate concentration,

1/ Vm = 3.0574
Vm = 0.3271
1/ K1. Vm = 0.0032
1/ Km(3.0574) = 0.0032
Km = 102mol/L
Solution 2.17
V= 5L
Cso = 100 mmol/L
F = 1 L/hr
Cs = 10m mol/L
a) F (s0 – FCs = rp V
1(100-10) = rp (5)
Rp = 18 m mol/ L.min
b) Find rp for each F and s
Equation obtained y= 0.0391x + 0.1641
M= 1/Vmax = 0.0391
Vmax= 25.57 m mol/L.min
Km/ Vm = 0.1641
Km= 4.197
Solution 2.18
[SO]1 = 0.1 mol/L [S0]2 = 0.3mol/L [E0] = 0.05 mol/L

[ ][ ]
[ ]
[ ][ ]
[ ]
[ ][ ]
[ ]
[ ][ ]
[ ]
[ ][ ]
[ ]
[ ][ ]
[ ]
[ ]
V1 = = k5[ES1]
=3.5 [ES1] ---

[ ]
V2 = = k6[ES2]
=2.8 [ES2] ---
[E0] = [E] + [ES1] + [ES2]

[ ][ ]
[E0] = [E] + [ES1] +
[ ]
[E0] = [ES1] + [E] (1+ )
[ ] [ ]
[E0] = [ES1] + [ ]
(1+ )
[ ]
[E0] = [ES1] {1 + [ ]
(1+ )}
[ ]
[ES1] = [ ]
( )
[ ]
[ ]
Vmt = [S1]0 – [S2] + K – ln [ ]
3.5 [ES1] t = 0.1 – [S1] +0.0714 ln [ ]
3.5 [ES1] t + [S1] = 0.1 +0.0714 ln 0.1 - 0.0714 ln [S1]
3.5 [ES1] t + [S1] + 0.0644 = -0.0714 ln [S1]

[ ] [ ]
ln[S1] =
[ ] [ ]
[S1] = ---
[E0] = [E] + [ES1] + [ES2]

[ ][ ]
[E0] = [E] + + [ES2]
[ ]
[E0] = [E] (1+ )+ [ES2]
[ ] [ ]
[E0] = [ ]
(1+ )+ [ES2]
[ ]
[E0] = [E2] [ [ ]
(1+ )+ 1]
[ ]
Vmt = [S1]0 – [S2] + KMln [ ]
2.8[ES1] t = 0.3 – [S2] + 0.2207ln [ ]
2.8[ES1] t + [S2] = 0.3 + 0.2207ln 0.3 – 0.2207ln [S2]
2.8[ES1] t + [S2] – 0.0343 = – 0.2207ln [S2]
[ ] [ ]–
ln[S2] = –
[ ] [ ]–
[S2] = e –
---
As [S1] increases, [ES1] also increases as in eq.3. [P1] also increases as in eq.1. This also occurs in
[S2]. As [S1] increases, [ES1] also increases as in eq.4. [P2] also increases as in eq.2
Solution 2.19
Data :
s s/v
6.7 22.33333
3.5 14
1.7 10.625
Langmuir Plot
25
y = 2.3722x + 6.2429
20
15
10
0
0 1 2 3 4 5 6 7 8
1/Vm = 2.3722
Vmax = 0.4215 mumol/L.min
Km/Vm = 6.2429
Km = 6.2429(0.4215)
=2.63mumol/L
Solution 2.20
a. Write the kinetic model.
Since the Michaelis constant KM is not affected by the presence of the inhibitor (which has shown on
the given table); then this enzyme reaction is noncompetitive inhibition reaction.
Kinetic Model:
E  S   ES
k 1, k 2
E  I   EI
k 3, k 4
EI  S   EIS
k 5, k 6
ES  I   ESI
k 7 ,k 8
ES 
k9
EP
b. Derive the rate equation. State the assumptions.
Assumptions:
 The dissociation constant for the first equilibrium reaction is the same as that of the third
equilibrium reaction.
 The dissociation constant for the second equilibrium reaction is the same as that of the
fourth equilibrium reaction.
The two equilibrium reactions,
k2 k
 K S  6  K IS
k1 k5
k4 k
 K I  8  K SI
k3 k7
If the slower reaction, the product formation step, determines the rate of reaction according to
Michaelis-Menten assumption, the rate can be expressed as:
rP  k 9 [ ES ] (1)
The enzyme balance gives
[ E0 ]  [ E ]  [ ES ]  [ EI ]  [ ESI ] (2)
Divide (1) by (2),
rP k 9 [ ES ]
 (3)
[ E0 ] [ E ]  [ ES ]  [ EI ]  [ ESI ]
Applied Law of mass action,
K 2 [ E ][S ] [ E ][S ]
Ks    [ ES ]  (4)
K1 [ ES ] KS
K 4 [ E ][ I ] [ E ][ I ]
KI    [ EI ]  (5)
K3 [ EI ] KI
k8 [ ES ][ I ] [ ES ][ I ]
KI    [ ESI ]  (6)
k7 [ ESI ] KI
Substitute (4), (5), (6) into (3),
[ E ][ S ]
k9
rP KS

[ E0 ] [ E ][ S ] [ E ][ I ] [ ES ][ I ]
[E]   
KS KI KI
[ E ][ S ]
k9
rP KS

[ E0 ] [ E ][ S ] [ E ][ I ] [ E ][ S ][ I ]
[E]   
KS KI KS KI
Eliminate [E],
[S ]
rP KS

[ E0 ]k 9 [ S ] [ I ] [ S ][ I ]
1  
KS KI KS KI
Substitute rPmax  [ E0 ]k 9
[S ]
rP KS

rPmax [ S ] [ I ] [ S ][ I ]
1  
KS KI KS KI
Multiply numerator and denominator by Ks,
rP [S ]

rPmax K [ I ] [ S ][ I ]
K S  [S ]  S 
KI KI
Rearranging,
rP [S ]

rPmax K [I ] [ S ][ I ]
KS  S  [S ] 
KI KI
rP [S ]

rPmax [I ] [I ]
K S (1  )  [ S ](1  )
KI KI

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BK10110302 V.

PRASARNTH RAAJ VEERA RAO – BIOPROCESS

Total volume = 44 + 5 + 1= 50ml

From the graph the equation obtained

m= 0.033 ɱ mol / ml . min

a) Activity of the β- glucosidase

0.033 x 50= 1.65 mumol / min

= 165 units/mg protein

ii) = 1.65 mumol/min

= 1.65 units/ml of enzyme

b) Initial rate of reaction 0.033 mumol/ mL.min

The rate of product formation.

Since the enzyme is preferred,

Make E as the subject,

Since forward reaction = backward reaction.

(a) E+S k1 (ES)1

[E0] = [E] + [ES] + [ES]2

k2 [ES]1 = [Eo] [S] – [ES]1 [S]

[ES]1 ( k2 + [S] ) = [E0] [S] – [ES]2 [S]

[ES]1 = [E0] [S] – [ES]2 [S]

k4 [ES]2 = [E0] [S] – [ES]2 [S]

[ES]2 ( k2 k4 + k4 [S] + [S] )= [E0] [S]

[ES]2 = [E0] [S]

V= d [P] = k5 [E0] [S]

The rate of product formation.

Since the enzyme is preferred,

Make E as the subject,

Since forward reaction = backward reaction.

The rate of product formation,

Since the enzyme is preferred,

Lineweaver- Burk Plot

Equation obtained y= 0.0172 x + 3.6342

y-intercept = 3.6342= 1/ V max

Equation obtained y= 3.3133x + 0.0191

y-intercept= km/Vm = 0.0191

Equation obtained y= -0.0043x + 0.2645

Non-Linear Regression Procedure

From the graph, Vm=0.2

½ Vmax = 0.1, Km=0.0032

Data for Graph plot :

Non-Linear Regression Plot

0.05 y = 3.3133x + 0.0191

Lineweaver Burk Plot

Non-Linear Regression Procedure

Rate of product formation

Substitute equation into

Dividing with the value of

a) FCs0 - FCs + rSv = V

For Batch reactor F=0

b) Equation obtained y = 6.3852x + 59.571

c) FCs0 - FCs + rSv = 0

( FCSo - FCs ) (Km + Cs) = Vmax CsV

(0.0001 (300)(200) + 0.0001(300)Cs – 0.001(200)Cs – 0.001Cs2 = 100 (0.0003)Cs )

Cso = 3.4 x 10 -4 mol/L

Cs = 0.9 x 3.4 x 10-4

= 3.06 x 10-4 mol/L

( 3.4x 10-4 – 3.06 x 10-4) = Vmax (3.06 x 10-4)

6.8 x 10-6 = Vmax ( 0.03)

b) 6.8 x 10-6 x 15 = 1.02 x 10-4 mol/L

CSTR @ Stead State

b) Plug - Flow @ Stead State

0.03 ln + (10 - 0.5 ) = 780t

CSTR @ Steady State