Exploring the Molecules of Life:  Enzymes

Enzymes are nature's catalysts, as they are compounds that enhance the rate (speed) of biochemical reactions.  The enzyme lysozyme is given below.  Look at its structure and decide on what the molecule is composed of: carbohydrates, lipids, or proteins.  Zoom in by holding the shift key down and click and move the mouse.  Hydrogens are not shown!

Using the right click, under options turn on the disulfide linkages (note: close mouse instructions window) and the hydrogen bonds.   By now you have figured out that enzymes are proteins.  Lysozyme is an enzyme that destroys the cell walls of bacteria by hydrolyzing the polysaccharide component of the cell wall.  Lysozyme is found in the mucosal membranes that line the human nasal cavity and tear ducts. 

The table summarizes a number of enzymes, the reactions they catalyze, the function of this reaction, and how the rate of the reaction is influenced by the enzyme.

Enzyme Reaction catalyzed Function of reaction Rate*
catalase 2H2O2    2H2O + O2 removes toxic H2O2 from cell 1015
carbonic anhydrase CO2 + H2 H2CO3 hydrates CO2 gas for transport 107
invertase sucrose + water  fructose + glucose breakdown of sugar --
pepsin proteins polypeptides digestion of proteins in stomach --
trypsin polypeptides amino acids further digestion of proteins in small intestines --

 * rate of enzyme catalyzed compared to rate uncatalyzed

Enzymes are helpful in DNA replication and protein synthesis.  They help by breaking the hydrogen bonds between the bases and allowing the strand to duplicate itself.

Enzymes are used according to the bodies need for them.  There are enzymes that aid in blood clotting, and those that aid in digestion, and even those within the cell that are needed for specific reactions.

Enzymes are usually given names based on the reactions they catalyze or the compound they are reacting on.  The names of most enzymes end in "-ase."  Some enzymes which have been known longer have older names that don't end in "ase."  For example: pepsin, trypsin, and chrymotrypsin which are all enzymes localized in the digestive tract.

There are six major groups of enzyme classification:

1. Oxidoreductases- enhance the rate of oxidations and reductions

2. Transferases- enhance the rate of the transfer of groups of atoms (CH3, CH3CO, or NH2) from one molecule to another.

3. Hydrolases- enhance the rate of hydrolysis reactions

4. Lyases- enhance the rate of the addition of one group to a double bond  or removal of two groups from nearby atoms to create a double bond.

5. Isomerases- enhance the rate of isomerization reactions.

6. Ligases or synthetases- enhance the rate of the joining of two molecules.

Click here for more information on the classification of enzymes.

Consider the graph of relative reactivity of an enzyme reaction against temperature.  How would you describe the graph?  Is it the same shape as a typical chemical reaction?

Temperature affects the activity of enzymes because it changes the three-dimensional structure of the enzyme.  In an uncatalyzed reaction the rate increases in direct proportion to the temperature. 

But, in catalyzed reactions the effect of temperature is quite different.  Starting at a low temperature, and increasing the temperature initially causes the rate to increase.  However, once the peak temperature has been reached the rate of the reaction begins to decrease.  After a small temperature increase above the peak, the rate decrease can be increased again by lowering the temperature, because over a narrow temperature range, changes in activity can be reversed.  Though, at a high temperature above the peak, there is a point where the enzyme becomes inactive.

Now consider a graph of relative reactivity of an enzyme reaction against pH.   How would you describe the graph?

Each enzyme functions the best at a certain pH.  Within a narrow pH range enzyme activity can be reversible.  However, high pH values (acidic or basic) can cause the enzyme to be completely inactive and some cannot be restored by going back to the peak pH.  This behavior is due to the acid-base groups of the amino acids contained in the enzyme.

To understand how an enzyme functions, go to the tutorial on catalase at the Online Macromolecule Museum.

Enzyme Kinetics

An enzyme (E) particaptes in a biochemical reaction; however, it is not consumed in the reaction.  The substance that reacts with the enzyme is called the substrate (S).  Here is a simple picture of what is going on as the substrate binds to the enzyme (ES) and the enzyme converts the substrate to a product (P).

E   +   S   =  ES  =   E   +   P

As the reaction proceeds, what do you notice about the enzyme?

From simple chemical kinetics, one might assume that the rate of substrate consumption would be given by:  Rate = k(E)(S) where k is a rate constant for the reaction.  If the enzyme concentration is constant then the equation becomes:  Rate = k'(S) which is the equation of a straight line.  Let's look at how this compares to real behavior.

How would you describe the behavior of an enzyme-catalyzed system compared to the reaction of a typical chemical system?

In the reaction above, the enzyme, E, is recovered to react again.  At any time during the reaction,  Eo = E + ES, where Eo is the initial enzyme concentration.  The rate of this reaction can be studied and measured.  Enzymes are known to follow Michaelis-Menten kinetics which are different from typical chemical reaction kinetics and governed by the equation below.

where V is the rate of the reaction, Vmax is the maximum rate (or kcatEo where kcat is a constant that depends on the enzyme and Eo is the initial enzyme concentration), Km is the Michaelis constant for the enzyme, and S is the substrate concentration.  Let's see how the graph of rate against substrate concentration behaves for an enzyme-catalyzed system. 

Click here to go to an interactive Excel spreadsheet that will allow you to make changes in the variables in the equation above and see how the data is influenced.  Use this interactive spreadsheet to address the following questions.

1.    How would you describe the curve generated by a plot of rate against substrate concentration?  How does its shape change if the variables are adjusted?

2.    How can you increase the value of Vmax?

3.    How does increasing the value of the Michaelis constant, Km, influence the graph?

4.    If you substitute  kcatEo for Vmax in the Michaelis-Menten equation above, how is the rate, V, and the initial enzyme concentration, Eo, related assuming the substrate concentration, S, is constant?

5.    If Km = (S), what does the rate, V, equal?

The curve you have just explored is a rectangular hyperbola.  It is not one of the common curves that are fit by regressions on graphing calculators or spreadsheets.  From the days of treating data by hand, this curve can be put into a linear form by transforming the data.  The most common plot is 1/V against 1/(S) and if you click on the tab labeled inverse plot at the bottom left of the Excel worksheet, you will see the transformed data plotted.  This plot can be changed by going back to the direct plot tab and changing the kcat, Eo, or Km .  

6.    Here is some data for glucose-6-phosphate converted to fructose-6-phospate by the enzyme phosphoglucose isomerse.  Using graph paper, a graphing calculator, or Excel plot the two types of graphs and determine  Vmax and Km.  The concentration units are  micromolar or 10-6 molar.  Need graphing calculator or Excel help - click here.

Substrate concentration (micromolar) Reaction rate (micromolar/minute)
0.08 0.15
0.12 0.21
0.54 0.70
1.23 1.1
1.82 1.3
2.72 1.5
4.94 1.7
10.00 1.8

From: http://biology.csusb.edu/300kinetics.html

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