GENERAL BIOLOGY LAB 1 (BSC1010L)

The active site determines the specificity of every enzyme. Specificity can result
from the charge, shape, and hydrophobic/hydrophilic characteristics of the enzyme and
substrate molecules. Only reactants that match the geometric shape of the active site can
bind to the enzyme. This specificity is also known as the “lock and key model” where
the substrate (the key) fits into the active site (the lock) of an enzyme. However, as
mentioned previously, the interaction between a substrate and the enzyme’s active site is
not static. When a substrate binds to the enzyme, the active site is reshaped by the
interactions of the enzyme’s amino acid side chains with the substrate molecule. This
protein remodeling enhances the overall binding of the reactant to the active site,
increasing catalytic action. The ability of enzymes to mould their shape to enhance the fit
of substrate molecules is known as the “induced fit model.”
There are thousands of different enzyme types, each with a specific set of
conditions at which it works best, i.e., its optimal conditions. An enzyme’s optimal
conditions often reflect the environment(s) of the organism(s) in which it is found. For
instance, the optimum temperature for enzymes present in Thermophilus aquaticus, an
extremophilic bacterium that inhabits hot springs, is about 70ºC. In contrast, peroxidase,
an enzyme present at high concentrations in turnips, horseradish roots and potatoes,
works best at temperatures around 45ºC.
Enzymatic activity is affected by multiple factors, including pH, substrate
concentration, salt concentration, as well as the presence of inhibitors, activators and
cofactors. In this lab, you will examine the effect of temperature on enzymatic reactions.
Temperature affects the rate at which substrate and enzyme molecules collide. At
temperatures greater than the optimal, the active site denatures (i.e. changes shape),
decreasing or preventing substrate binding. Consequently, product formation is either
reduced or completely arrested. At the other end of the spectrum, low temperatures
decrease the movement of molecules, resulting in less contact between enzymes and
substrates, which slows down the frequency and rate of reaction, and ultimately
diminishes product formation. Although the effect of temperatures outside of the optimal
range on substrate catalysis is the same, the mechanism through which enzyme activity is
reduced, differs.
The objective of the current lab is two-fold; (1) to examine how variations in
temperature affect the activity of the enzyme amylase and (2) to determine the optimal
temperature for amylase from two different sources (fungal and human).
Amylase catabolizes starch polymers (a storage polysaccharide) into smaller
subunits (monomers = saccharides) including maltriose, maltose and short
oligosaccharides comprised of 2-20 monosaccharide units (Fig. 3). Most organisms use
these saccharides as a food source and to store energy (Fig. 4). Both starch and amylase
are important commercially in the production of syrups and other food products, as well
as for fermentation and brewing processes.
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