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Bio200 POGIL Cell Biology Activity 2 – Enzymes Schivell

How do enzymes work?


NOTES:
MODEL 1: - His 12, Lys41, and His119 are key parts of the
active site (shown in the gray box and expanded
Substrate below), which is a small part of the enzyme.

- The substrate is RNA (ribonucleic acid).

Enzyme - Phosphorus (P) is most stable with 5 bonds and has


about the same electronegativity as C and H.

- This enzyme "catalyzes" the reaction shown.

REACTION MECHANISM

1. 2. 3.

6. 5. 4.

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Bio200 POGIL Cell Biology Activity 2 – Enzymes Schivell
CRITICAL THINKING QUESTIONS:

1. What biological macromolecule is the enzyme made of? __________________________

2. Compare panels 1 and 6 of the reaction mechanism (these contain the initial and final states
of the RNA substrate). Describe in fewer than 5 words what has changed about the RNA.

For the following questions, it is a good idea to circle or annotate the differences you see on
the model itself, or to draw arrows indicating formation of new bonds or movements of
atoms.

3. In panel 2 compared to 1, what has changed for the... (be as brief but specific as possible)

... substrate? __________________________________________________________

... enzyme? ___________________________________________________________

4. In panel 3 compared to 2, what has changed for the...

... substrate? __________________________________________________________

... enzyme? ___________________________________________________________

5. a. In panel 4 compared to 3, what has changed for the...

... substrate? __________________________________________________________

... enzyme? ___________________________________________________________

b. What new molecule is introduced in panel 4? ____________

6. In panel 5 compared to 4, what has changed for the...

... substrate? __________________________________________________________

... enzyme? ___________________________________________________________

7. In panel 6 compared to 5, what has changed for the...

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Bio200 POGIL Cell Biology Activity 2 – Enzymes Schivell
... substrate? __________________________________________________________

... enzyme? ___________________________________________________________

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Bio200 POGIL Cell Biology Activity 2 – Enzymes Schivell
8. In panel 1, find two different types of molecular interactions that help attract and hold the
substrate in the enzyme's active site. (Choose from hydrophobic interactions, covalent bonds,
hydrogen bonds, and ionic bonds.) Write the name of each interaction and draw an example
of the atoms/groups of atoms involved in the boxes below:

9. a. In panel 2, which atom has an unusual number of bonds? _________________

b. Does the substrate in panel 2 have a higher or lower G than in panel 1? _____________

10. Consider the enzyme's role in the reaction mechanism:

a. Describe the "job" of the two histidine (His) R-groups. Protonation site and
deprotonation site

b. Describe the "job" of the lysine (Lys) R-group. Ionic bond

11. a. The R-groups of the active site are in the same conformation in panels 1 and 6. Can the
enzyme catalyze another reaction after it completes this one? Why? Yes, same structure as
before.

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Bio200 POGIL Cell Biology Activity 2 – Enzymes Schivell

MODEL 2:
The wild-type enzyme and the rate at which the reaction
proceeds in its presence is shown to the right. Below are
three different enzyme mutants. For each, one aspect of
the active site's structure is slightly changed. The mutant
enzymes are mixed with RNA in solution. The reaction
rates are shown underneath each image.

WT = 1000 rxns/sec

(His12 shifted "up"


slightly) (Lys41 is now Met41) (His119 is now Val119)

Mut1 = 10 rxns/sec Mut2 = 1 rxn/hour Mut3 = 1 rxn/millenium

12. How does reaction speed change when the active site is changed?
________slows_____________

13. a. Is the relative position of a specific R-group within an active site important?
___yes______

b. Which mutant helps to answer this question? ___mut1 – further away____

14. When the transfer of a proton between the enzyme and


substrate is prevented, is the reaction rate changed slightly or dramatically? __dramatically –
see mut 3________

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Bio200 POGIL Cell Biology Activity 2 – Enzymes Schivell
15. Even if an R-group (that is part of the active site) does not normally accept or donate
protons or other atoms, or form covalent bonds with the substrate, can it be important?
Explain. Ionic bonds between the lys and the substrate

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Bio200 POGIL Cell Biology Activity 2 – Enzymes Schivell
On your own:
The drawing shows the first 2 steps of a 5 step
A
reaction mechanism. The enzyme's peptide backbone
is a solid line (except for some atoms shown in detail).
The two arrows (A and B) point to the two ends of the
enzyme. The two R-groups of the active site, CYS and
HIS, are shown in detail. In the first frame, the enzyme's
two substrates are shown, NAD+ and a second
substrate in the dotted line.

1. What type of biological macromolecule


is the second substrate (ignore phosphate) carb

2. Which end (A or B) is the amino


terminus of the enzyme? _____ B

3. Which of the following R-groups would be best Step 1


suited to be in the "X" location shown in the first
diagram? (Circle ONE)

4. Which terms describe the reaction catalyzed


by this enzyme?
(Circle ALL that apply)

-endergonic - catabolic - spontaneous - ATP-driven

5. In Step 1, how does HIS participate in the reaction? Step 2


(1 sentence or less)

6. What is transferred between the two substrates


during Step 2?

7. Can the enzyme catalyze another reaction after Step 2? _______

9. Using the enzyme to catalyze the reaction changes which


of the following? (Circle ALL that apply)
- ∆G - ∆S - whether it is spontaneous − ∆G‡ (EA)

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Bio200 POGIL Cell Biology Activity 2 – Enzymes Schivell
10. If CYS were changed to Valine (see below), how would
the rate of catalysis change? Explain in 1-2 sentences.