Enzyme Activity As A Function Of Substrate Concentration Biology Essay

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The effect of concentration on enzyme activity. It can be noted that both enzyme solution used had different results. Enzyme activity was greater when using the chicken liver rather than the cow liver.

Generally the enzyme activity increases as the drops of the enzyme (concentration) increases.

There was a significant increase when six drops of enzyme solution (chicken liver) was added. The height of the bubble rose to 4 cm. there was a steady increase between when 9-12 drops solution was added. A sharp increase occurred when 15 drops of enzyme solution was added. The height was recorded as 7.5 cm. There was no significant increase onwards

When enzyme solution (cow's liver) was added there was no significant increase in the first set of drops. The height was recorded to be 3.5 cm when 3 drops of enzyme solution was added. There was an increase from 3.5 to 4cm when 6 drops of solution was added. There were no further significant increases un adding the enzyme solution.

Graph 2 shows the effect of temperature on enzyme activity. For this experiment an enzyme solution of cow liver was used. There was an increase in bubble height when the solution was placed at 25 C. There was a further increase in the height when the solution was placed at 50 C. The height was recorded as 4.2 cm. However at 70 C, there was no change. There was no evidence of bubbles suggesting that the enzyme had been denatured.

EXPLANATION

Generally enzyme activity increases with increasing temperature. However in this experiment there was an increase then a sudden drop in enzymatic activity. Enzymes require certain conditions to be effective in their functioning. Some require certain temperatures to function. A reason behind the denaturing of the enzyme at 70 C is that the temperature was too high for the enzyme to function and it denatured due to this.

Factors Affecting Enzyme Activity

Knowledge of basic enzyme kinetic theory is important in enzyme analysis in order both to understand the basic enzymatic mechanism and to select a method for enzyme analysis. The conditions selected to measure the activity of an enzyme would not be the same as those selected to measure the concentration of its substrate. Several factors affect the rate at which enzymatic reactions proceed - temperature, pH, enzyme concentration, substrate concentration, and the presence of any inhibitors or activators.

Temperature Effects

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Like most chemical reactions, the rate of an enzyme-catalyzed reaction increases as the temperature is raised. A ten degree Centigrade rise in temperature will increase the activity of most enzymes by 50 to 100%. Variations in reaction temperature as small as 1 or 2 degrees may introduce changes of 10 to 20% in the results. In the case of enzymatic reactions, this is complicated by the fact that many enzymes are adversely affected by high temperatures. As shown in Figure 13, the reaction rate increases with temperature to a maximum level, then abruptly declines with further increase of temperature. Because most animal enzymes rapidly become denatured at temperatures above 40°C, most enzyme determinations are carried out somewhat below that temperature.

Over a period of time, enzymes will be deactivated at even moderate temperatures. Storage of enzymes at 5°C or below is generally the most suitable. Some enzymes lose their activity when frozen.

Effects of pH

Enzymes are affected by changes in pH. The most favorable pH value - the point where the enzyme is most active - is known as the optimum pH. This is graphically illustrated in Figure 14.

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Extremely high or low pH values generally result in complete loss of activity for most enzymes. pH is also a factor in the stability of enzymes. As with activity, for each enzyme there is also a region of pH optimal stability.

The optimum pH value will vary greatly from one enzyme to another

Enzyme Concentration

In order to study the effect of increasing the enzyme concentration upon the reaction rate, the substrate must be present in an excess amount; i.e., the reaction must be independent of the substrate concentration. Any change in the amount of product formed over a specified period of time will be dependent upon the level of enzyme present. Graphically this can be represented as:

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These reactions are said to be "zero order" because the rates are independent of substrate concentration, and are equal to some constant k. The formation of product proceeds at a rate which is linear with time. The addition of more substrate does not serve to increase the rate. In zero order kinetics, allowing the assay to run for double time results in double the amount of product.

The amount of enzyme present in a reaction is measured by the activity it catalyzes. The relationship between activity and concentration is affected by many factors such as temperature, pH, etc. An enzyme assay must be designed so that the observed activity is proportional to the amount of enzyme present in order that the enzyme concentration is the only limiting factor. It is satisfied only when the reaction is zero order.

Enzyme activity is generally greatest when substrate concentration is unlimiting.

Effects of Inhibitors on Enzyme Activity

Enzyme inhibitors are substances which alter the catalytic action of the enzyme and consequently slow down, or in some cases, stop catalysis. There are three common types of enzyme inhibition - competitive, non-competitive and substrate inhibition.

Most theories concerning inhibition mechanisms are based on the existence of the enzyme-substrate complex ES. Competitive inhibition occurs when the substrate and a substance resembling the substrate are both added to the enzyme. A theory called the "lock-key theory" of enzyme catalysts can be used to explain why inhibition occurs.

The lock and key theory utilizes the concept of an "active site." The concept holds that one particular portion of the enzyme surface has a strong affinity for the substrate. The substrate is held in such a way that its conversion to the reaction products is more favorable. If we consider the enzyme as the lock and the substrate the key - the key is inserted in the lock, is turned, and the door is opened and the reaction proceeds. However, when an inhibitor which resembles the substrate is present, it will compete with the substrate for the position in the enzyme lock. When the inhibitor wins, it gains the lock position but is unable to open the lock. Hence, the observed reaction is slowed down because some of the available enzyme sites are occupied by the inhibitor. If a dissimilar substance which does not fit the site is present, the enzyme rejects it, accepts the substrate, and the reaction proceeds normally.

ACTIVE SITE

This is the part of an enzyme or antibody where the chemical reaction occurs

SUBSTRATE

In biochemistry, a substrate is a molecule upon which an enzyme acts. Enzymes catalyze chemical reactions involving the substrate(s). In the case of a single substrate, the substrate binds with the enzyme active site, and an enzyme-substrate complex is formed. The substrate is transformed into one or more products, which are then released from the active site

ENZYME SUBSTRATE COMPLEX

This is when the substrate binds reversibly to the enzyme forming a complex

ANABOLISM

Anabolism, or biosynthesis, is the process by which living organisms synthesize complex molecules of life from simpler ones. Anabolism, together with catabolism, are the two series of chemical processes in cells that are, together, called metabolism. Anabolic reactions are divergent processes. That is, relatively few types of raw materials are used to synthesize a wide variety of end products. This results in an increase in cellular size or complexity-or both.

Anabolic processes produce peptides, proteins, polysaccharides, lipids, and nucleic acids. These molecules comprise all the materials of living cells, such as membranes and chromosomes, as well as the specialized products of specific types of cells, such as enzymes, antibodies, hormones, and neurotransmitters.

CATBOLISM

Catabolism, the opposite of anabolism, produces smaller molecules used by the cell to synthesize larger molecules, as will be described below. Thus, in contrast to the divergent reactions of anabolism, catabolism is a convergent process, in which many different types of molecules are broken down into relatively few types of end products.

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