Factors Affecting Enzyme Reactions

• Factors like temperature, pH and concentration can effect how well enzymes work.
• Like any chemical reaction a higher temperature initially increases the rate of an enzyme-controlled reaction.

  • The enzyme and substrate molecules have more kinetic energy, move faster and are more likely to collide.
  • This leads to a faster rate of reaction.
• Heating to high temperatures (beyond the optimum) will break the bonds that hold the enzyme together and the active site will lose its shape.

  • the enzyme has been denatured irreversibly and will not go back into its original form.
  • The substrate will not fit into the active site any more,.
  • The enzyme can no longer catalyse the reaction so it stops.
• Enzymes work fastest at their ‘optimum temperature’.

  • In the human body, this optimum temperature is about 37⁰C which is our normal body temperature.
• Like this diagram... (1st part)

  
• Like this diagram... (2nd part)

  
• Graph showing the effect of temperature on enzyme activity.

  
• As we increase the temperature, the activity of the enzyme increases (the reaction gets faster). That's because as the temperature increases, the enzyme and substrate are moving faster, so there are more collisions per second, between the substrate and the active site. Like this...

  
• At a certain temperature, the enzyme is working at the fastest possible rate. That's called the optimum temperature. At this point, there is the maximum frequency of successful collisions between the substrate and the active site. Like this...

  
• As we increase the temperature past the optimum, then the activity of the enzyme rapidly decreases to zero (the enzymes stop working).

  like this...

  
• At high temperature, the enzyme molecule vibrates and the shape of the active site changes. Like this...

  
• Now the substrate no longer fits perfectly into the active site. The active site is denatured. Like this...

  The enzyme can no longer catalyse the reaction.

  
• The enzyme has an optimum PH, where the activity is maximum. Like this...

  
• If we make the PH more acidic or more alkaline then the activity drops to zero. That's because the active site denatures if the conditions are too acidic or too alkaline. Like this...

  
• The pH also has an effect on enzymes, if it is too high or too low it interferes with the enzyme.

  • The optimum pH for most human enzymes is pH 7.
  • Enzymes produced in acidic conditions, such as the stomach, have a lower optimum pH (pH 2).
  • Enzymes produced in alkaline conditions, such as the duodenum, have a higher optimum pH (pH 8 or 9).
• If the pH is too far above or too far below the optimum, the bonds that hold the amino acid chain together to make up the protein can be disrupted or broken.

  • This changes the shape of the active site, so the substrate can no longer fit into it, reducing the rate of activity.
• Moving too far away from the optimum pH will cause the enzyme to denature and the reaction it is catalysing will stop. Like this...

  
• Graph showing the effect of pH on the rate of activity for an enzyme from the duodenum.

  
• The greater the substrate concentration, the greater the enzyme activity and the higher the rate of reaction:
• • As the number of substrate molecules increases, the likelihood of enzyme-substrate complex formation increases.
  • If the enzyme concentration remains fixed but the amount of substrate is increased past a certain point, however, all available active sites eventually become saturated and any further increase in substrate concentration will not increase the reaction rate.
  • When the active sites of the enzymes are all full, any substrate molecules that are added have nowhere to bind in order to form an enzyme-substrate complex.
• The effect of substrate concentration on the rate of an enzyme-catalysed reaction.

  
• Practical - The effect of pH on the rate of reaction of amylase:

  • Amylase is an enzyme that breaks down starch (a polysaccharide of glucose) into maltose (a disaccharide of glucose).
  • The effect of different pH levels on the activity of amylase can be investigated.
  • The rate of reaction can be easily monitored by detecting the presence of starch.
  • Starch can be detected using iodine solution.
  • If starch is present, the iodine solution will change from a brown/orange colour to blue-black.
• Method: (Part 1)

  • Add a drop of iodine to each of the wells of a spotting tile.
  • Use a syringe to place 2 cm3 of amylase into a test tube.
  • Add 1cm3 of buffer solution (at pH 2) to the test tube using a syringe.
  • Use another test tube to add 2 cm3 of starch solution to the amylase and buffer solution, start the stopwatch whilst mixing using a pipette.
• Method: (Part 2)

  • Every 10 seconds, transfer a droplet of the solution to a new well of iodine solution (which should turn blue-black).
  • Repeat this transfer process every 10 seconds until the iodine solution stops turning blue-black (this means the amylase has broken down all the starch).
  • Record the time taken for the reaction to be completed.
  • Repeat the investigation with buffers at different pH values (ranging from pH 3.0 to pH 13.0).
• Results and Analysis: (part 1)

  • At the optimum pH, the iodine remained orange-brown within the shortest amount of time.
  • This is because the enzyme is working at its fastest rate and has digested all the starch.
  • At higher or lower pH's (above or below the optimum) the iodine took a longer time to stop turning blue-black or continued to turn blue-black for the entire investigation.
  • This is because on either side of the optimum pH, the enzymes are starting to become denatured and as a result are unable to bind with the starch or break it down.
• Results and Analysis: (part 2)

  The time taken for the disappearance of starch is not the rate of reaction.

  • It gives us an indication of the rate but it is actually the inverse of the rate.
  • The shorter the time taken, the greater the rate of the reaction.
  • In this case, you can still calculate the rate of reaction by using the following formula:

  Rate = 1 ÷ Time

  • Once the rates of starch breakdown at different pH has been determined the results can be represented on a graph.
• A graph showing the optimum pH for an amylase in the breakdown of starch. Like this...

  
• Rate calculations for enzyme activity:

  • Rate calculations are important in determining how fast an enzyme is working (i.e. the rate of reaction)
  • To perform a rate calculation, use the following formula:


  Rate = Change ÷ Time

  Rate = Amount of substrate used or product formed ÷ Time