Spingospine 1-phosphate (SPP) has recently been discovered to be important for t

Question

Spingospine 1-phosphate (SPP) has recently been discovered to be important for the cell survival. The
synthesis of SPP from Spingospine and ATP is catalyzed by the enzymes Spingospine kinase. An
understanding of the kinetics of the Spingospine kinase reaction may be important in the development of
drugs to treat cancer. The velocity of Spingospine kinase reaction was measured in the presence and absence
of threospingospine, a stereoisomer of singspiel. The results are shown below. (15 pts)
[Spingospine](M)         Vo (mol/min)(No inhibitor)          Vo (mol/min) (With threospingospine)
2.5                                          32.3                                                     8.5
3.5                                          42                                                        11.5
5                                             48                                                        16.0
10                                           68                                                        28.0
20                                           87.7                                                     44.0
50                                           120                                                      74.0
Construct a Lineweaver –Burk plot to answer the following questions:
a) What are the Km value and Vmax in the presence and absence of the inhibitor?
b) What kind of an inhibitor is threospingospine? Explain.
c) Determine the Ki for the inhibitor ([I] = 5 M)

Explanation / Answer

For the given enzyme kinetics, calculate 1/s, and 1/Vo and plot the graph with 1/S on x-axis and 1/Vo on y-axis

Lineweaver-Burk plot

When there is no inhibitor:

1/Vmax = 0.0083, and Vmax = 1/0.0083 = 120.5 micro M/min

Km = 0.057 X 120.5 = 6.9 or approx. 7.0 micro M

In the presence of inhibitor:

1/Vmax = 0.0084, and Vmax = 1/0.0084 = 119 micro M/min

Km = 0.2733 X 119 = 32 micro M

b) This is a competetive inhibition. The two plots have the same or nearly same Vmax but differ in their Km values, which is characteristic of a competetive inhibition.

c) For the enzyme kinetics given, the value of KI =

1/s (M) 1/vo (micro mol/min) 1/vo 0.4 0.03096 0.117647 0.285714 0.02381 0.086957 0.2 0.020833 0.0625 0.1 0.014706 0.035714 0.05 0.011403 0.022727 0.02 0.008333 0.013514

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