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Bio Assignment 1

1) The document describes an enzymatic reaction that exhibits competitive inhibition. Plotting 1/V vs 1/S shows the reaction has a constant Vmax but increasing Km,app with higher inhibitor (I) concentrations. 2) Values for Km, Vmax, and Ki were determined from Lineweaver-Burk plots. Km was found to be 1.23 mM and Vmax was 6.34 mM/h. Ki was calculated to be 2.16 mM and 1.71 mM from two inhibitor concentrations, showing experimental error. 3) For an inhibitor concentration of 1.3 mM, the Ki was calculated to be 1.82 mM from the Michaelis-Ment

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41 views4 pages

Bio Assignment 1

1) The document describes an enzymatic reaction that exhibits competitive inhibition. Plotting 1/V vs 1/S shows the reaction has a constant Vmax but increasing Km,app with higher inhibitor (I) concentrations. 2) Values for Km, Vmax, and Ki were determined from Lineweaver-Burk plots. Km was found to be 1.23 mM and Vmax was 6.34 mM/h. Ki was calculated to be 2.16 mM and 1.71 mM from two inhibitor concentrations, showing experimental error. 3) For an inhibitor concentration of 1.3 mM, the Ki was calculated to be 1.82 mM from the Michaelis-Ment

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Swathi S
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1) (a) Plot 1/V vs 1/S for all cases to determine the type of inhibition

Vmax is the same, regardless of inhibition, but as I increases, –1/Km decreases (apparent), that is
an increased value of Km,app, resulting in a reduction in reaction rate. This is characteristic of
Competitive Inhibition

(b) Determine Vm, Km, KI


Vm, Km can be determined from the I = 0 case.@ I = 0 (1/V) = 0.15782 + 0.19404 (1/S)

1 Vm at  1 S   0 : 1 Vm  0.15782
Vm  6.34 mM h

1/ K m at  1/ Vm   0 : 0  0.15782  0.19404  1/ K m 


0.15782 1
 K m  1.23mM
0.19404 K m

KI must be determined from the I > 0 case

I  1.26 mM  '


  determine K m, app
I  1.95 mM 

I  1.26mM :  1 V   0.15917  0.3098  1 S 


0  0.15917  0.3098  1 K m,app 
0.15917 1
  K m,app  1.95 mM
0.3098 K m,app

I  1.95 mM :  1 V   0.15343  0.40383  1 S 


0  0.15343  0.40383  1 K m,app 
0.15343 1
  K m,app  2.63mM
0.40383 K m,app

  I 
K m,app  K m 1  
 KI 

 1.26mM 
I  1.26 mM : 1.95 mM  1.23mM 1  
 KI 
1.26
1.58  1 
KI
1.26
0.58   K I  2.16 mM
KI
 1.95mM 
I  1.95 mM : 2.63mM  1.23mM 1  
 KI 
1.95
2.14  1   K I  1.71mM
KI

We would expect that KI is the same at different concentrations of inhibitor. The differences in
the two values shown here are due to experimental error.

3) Plot 1/V versus 1/S at different inhibitor concentrations


Since the lines intercept at the same point on y-axis inhibition is competitive (Constant Vm,
increased Km).
For I = 0 , No inhibitor : From the intercept on y axis , 1/Vm = 0.2 and Vm = 5 mM/h
And from the intercept on X-axis, - 1/Km = -1.2 and Km = 0.83 mM
From 1/V versus 1/S plot for I = 1.3 mM and S0 = 0.50 mM V = 1.3 mM/h
Then, V = Vm S/ (Km(1+I/Ksı)) + S , 1.3 = 5(0.5)/ (0.83(1+1.3/Kı) + 0.5 )
Then Kı = 1.82 mM

E
H 2 O  CO2    HCO3  H 
4)
Plot 1/V versus 1/S (Lineweaver – Burk plot)
1 1 1

y – intercept = Vm k 2 E 0 , x – intercept = K m
1 40
  4  103
V  Co 2 
Hydration:

1 1 Vm  2.50  104 M sec 1


 4000M sec 
Vm r2  8.93 104 sec 1
1 4000
  100M 1  K m  0.01M
Km 40

1 163.15
  1.31 104
V  HCO3 

Dehydration:

1 V  7.61105 M sec 1
 1.31 104 M 1 sec  m
Vm r2  2.72 104 sec 1
1 1.31104 1
 M  K m  1.24 102 M
Km 163.15
5) Plot 1/V versus 1/S

a.) For E0 = 1.6 g/L


Km = 0.0246 mmol/ml at 30°C
Km = 0.0238 mmol/mlat 49.6°C

1 mmol
Vm   3.31
b.) 0.302 ml.min

c.) The inhibitor is competitive.


  I   I   0.6 mmol mL
K m,app  K m 1  
 KI  K m,app  0.052 mmol mL

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