Practice questions Spring 13: The hydration of CO2 is catalyzed by carbonic anhydrase. The data...

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Practice questions Spring 13:

The hydration of CO₂ is catalyzed by carbonic anhydrase. The data below was collected for this enzyme. Determine the K_m, v_max and k_catfor this enzyme. The concentration of the enzyme in the experiment was 3.4 X 10^-7 M. Is this a near prefect enzyme? Explain your answer.

[CO2] µM	Velocity µM/min
3	10.4
5	14.5
10	22.5
30	33.8
90	40.5

PLP is required for the enzyme ornithine aminotransferase to be functional. The enzyme was purified to homogeneity from a culture of cells grown in the presence of PLP. The activity of the enzyme was monitored as a function of time. Substrate was added and samples were withdrawn at times afterwards. Initially there is activity but after 2hours time the enzyme seems to have lost all activity. More substrate is added but no reaction occurs. What can you conclude about the role of PLP in this reaction?

The amino acid acyl-tRNA synthetases yield similar plots of 1/v versus 1/[substrate]. If the tRNA concentration is held constant and either ATP or the amino acid concentration is varied as the substrate and varying constant concentrations of the other substrate are added the line intersect on the x-axis and all have different slopes (see Figure 2 below). However if the t-RNA is added at varying constant concentrations then the lines all have the same slope (see Figure 2 below).
1. Does the binding of ATP and the amino acid occur in a Ping Pong or a single displacement? Explain your answer based on the kinetics data.
  1. If it is single displacement is it random or ordered
2. Does the binding of tRNA occur in a Ping Pong or a single displacement?
  1. If it is single displacement is it random or ordered

Figure 2:Kinetics graphs for Question 4

005_1ucgajo-l4u3ftdo.doc

David · Accepted Answer

Instructions: Graduate students must answer all questions for full credit
	1. The hydration of CO2 is catalyzed by carbonic anhydrase. The data below was collected for this enzyme. Determine the Km, vmax  and kcat for this enzyme.  The concentration of the enzyme in the experiment was 3.4 X 10-7 M. Is this a near prefect enzyme? Explain your answer.
[CO2] μM
	Velocity μM/min
	3
	10.4
	5
	14.5
	10
	22.5
	30
	33.8
	90
	40.5
	
	
Answer
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MichaelisMenten equation, 
V  VmaxS/(KM  S)
V  catEtS / ( M  S)
Vmax  catEt
cat incorporates the rate constants for 
all the reactions between 
ES and E  P 
in a multistep enzymatic process. 
kcat/
-
=+
=+
=
+
kK
k
k
KM ====
iscalledenzymeefficency
---
A graph between 1/V and 1/S can be drawn. The slope of the graph gives  Km/Vmax and intercept gives 1/Vmax.
Thus
Slope/intercept = Km
Km= 4.4592/0.0997
= 44.7261
Intercept =1/V max
V max= 1/0.0997
= 10.03
KCat= Vmax/[E]
= 10.03/3.4 X 10-7
= 2.95 x10^8
Enzyme efficiency = Kcat/Km
= 2.95 x10^8/44.

Practice questions Spring 13: The hydration of CO 2 is catalyzed by carbonic anhydrase. The data below was collected for this enzyme. Determine the K m , v max and k cat for this enzyme. The...

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