Mod 7 8

M8 – Properties of Protein
Physical Properties of Proteins

Proteins are colorless and usually tasteless. They are
homogenous and crystalline. Proteins range in shape. Some are
globular while some and fibrous. Globular proteins are spherical in
shape and are formed by folding and crumpling of protein chains.
Fibrous proteins are thread-like or ellipsoidal in shape and occur
mostly in animal muscles.
Proteins have large molecular weights and because of

their giant size, they exhibit many colloidal properties. Proteins are
amphoteric. They act as an acid and a base. Each protein has a
fixed value of the isoelectric point at which it will move in an electric
field.
The solubility of protein is influenced by pH. Solubility is lowest at an

isoelectric point and increases with increasing acidity or alkalinity.
Examples of proteins:
Albumin is a water-soluble, heat coagulable protein. It is present in egg white (egg albumin),
milk (lactalbumin), and serum. Egg albumin and lactalbumin are glycoconjugate proteins,
whereas serum albumin is a simple protein.
Casein is the chief protein of milk. It is a phosphoprotein. It is

insoluble in water but soluble in dilute acids and alkali. It is a non-heat coagulable
protein. It is rich in most of the essential amino acids. It contains traces of cysteine and
cystine.
Gelatin is insoluble in cold water but soluble in hot water. It is a non-

heat coagulable protein. It is derived from collagen. It is rich in
proline, hydroxyproline, glycine, and hydroxylysine but contains traces
of phenylalanine, tyrosine, cysteine, cystine, and methionine. It does
not contain tryptophan, hence it is a protein of low biological
value.
Denaturation
Denaturation is the loss of the secondary, tertiary, and quaternary
structures of a protein by a chemical or physical agent that leaves the
primary structure intact. The disruption of the primary protein
structure occurs with the hydrolysis of the peptide bonds to produce
free amino acids.
If these changes occur to a small extent, denaturation can be

reversed. For example, a denatured protein can be removed from a
urea solution and put it back into the water, it often reassumes its
secondary and tertiary structures. This process is called reversible
denaturation. In living cells, some denaturation caused by heat can
be reversed by chaperones. These proteins help a partially heat-
denatured protein to regain its native secondary, tertiary, and
quaternary structures. Some denaturation, however, is irreversible.
Factors that Cause Protein Denaturation:
1. Strong acids and bases
Changes in the pH can disrupt hydrogen bonds and salt bridges,

causing irreversible denaturation. Proteins are coagulated by strong
acids like concentrated HCl, H2SO4, and HNO3. Alkaloidal reagents such
as tannic acid and picric acid form insoluble compounds with protein.
It denatures protein irreversibly by disrupting the salt bridges and the
disulfide bonds.
2. Organic Solvents
Alcohol coagulates all types of protein except prolamines. Alcohol

irreversibly denatures by forming hydrogen bonds that compete with
the naturally occurring hydrogen bonds in the protein. At a
concentration of 70%, ethanol penetrates bacteria and kills them by
coagulating their proteins, whereas 95% of alcohol denatures only
surface proteins.
3. Reducing Agents
Reducing agents, such as 2-mercaptoethanol (HOCH2CH2SH), can break
the –S-S- disulfide bonds, reducing them to -SH groups. Reducing
agents also disrupts hydrogen bonds and hydrophobic interaction.
4. Salt Concentration
Changes in the salt concentration may result in the formation of a

precipitate (salting out). Salts affect both salt bridges and hydrogen
bonds.
5. Heavy Metals
Heavy metal salts like mercuric chloride or silver nitrate, and lead
precipitate protein. It also cleaves –SH bonds. It denatures protein
irreversibly by disrupting the salt bridges and the disulfide bonds.
6. Temperature Change
Heat cleaves hydrogen bonds, so boiling a protein solution destroys

the α-helical and β-pleated sheet structure.
7. Mechanical Stress
Stirring and grinding action may disrupt the delicate balance forces
required to maintain protein structure.
Biuret Test
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 It is the general test for the identification of
proteins.
 In the presence of an alkaline solution,
Cu2+ ion forms as a complex with the peptide
bonds due to the unshared electron pairs in
nitrogen, and the oxygen in the water.
 Once the complex has been formed, the
solution turns from blue to purple.
Ninhydrin Test
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 It is the test for the presence of α-amino acid
in proteins.
 Ninhydrin causes oxidative decarboxylation
and deamination of α-amino acids producing
an aldehyde, carbon dioxide, and ammonia.
 Ninhydrin is reduced to hydrindantin which
reacts with the liberated ammonia and
another molecule of ninhydrin.
Xanthoproteic Test
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 It is the test for the presence of aromatic
amino acids.
 Amino acids containing an aromatic nucleus
react with concentrated HNO3 to form a
yellow-colored complex on heating, it
changes to orange-red color when excess
NaOH is added.
Millon’s Test
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 It is the test for phenolic compounds.
 It is also used to detect the presence of
tyrosine – the only amino acid containing a
phenol group.
 Tyrosine is first nitrated by nitric acid in the
test solution, then the nitrated tyrosine
complexes mercury (I) and mercury (II) in the
solution to form either a red precipitate or a
red solution.
Hopkins-Cole Test
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 It is a test for the presence of tryptophan –
the only amino acid containing an indole
group.
 The indole reacts with glyoxylic acid in the
presence of a strong acid to form a violet
cyclic product.
Lead Sulfide Test
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 It is a test for the presence of cysteine or
cystine.
 The organic sulfur in cysteine or cystine is
released as inorganic S-8 ions which form
lead sulfide.
 The positive result is the formation of black
or brown precipitate
M10 Lesson 1 - Enzymes
A fundamental task of proteins is to act as enzymes—catalysts that

increase the rate of virtually all the chemical reactions within cells.
Although RNAs (ribozymes) are capable of catalyzing some reactions,
most biological reactions are catalyzed by proteins. In the absence of
enzymatic catalysis, most biochemical reactions are so slow that they
would not occur under the mild conditions of temperature and
pressure that are compatible with life.
Enzymes accelerate the rates of such reactions by well over a million-

fold, so reactions that would take years in the absence of catalysis
can occur in fractions of seconds if catalyzed by the appropriate
enzyme.
Enzymes are characterized by two fundamental properties.
 First, they increase the rate of chemical reactions without themselves

being consumed or permanently altered by the reaction.
 Second, they increase reaction rates without altering the chemical
equilibrium between reactants and products.
The binding of a substrate to the active site of an enzyme is a very
specific interaction. Active sites are clefts or grooves on the surface
of an enzyme, usually composed of amino acids from different parts of
the polypeptide chain that are brought together in the tertiary
structure of the folded protein. Substrates initially bind to the active
site by noncovalent interactions, including hydrogen bonds, ionic
bonds, and hydrophobic interactions. Once a substrate is bound to the
active site of an enzyme, multiple mechanisms can accelerate its
conversion to the product of the reaction.
The simplest model of enzyme-substrate interaction is the lock-and-

key model, in which the substrate fits precisely into the active site.
In many cases, however, the configurations of both the enzyme and
substrate are modified by substrate binding—a process called induced
fit. In such cases, the conformation of the substrate is altered so that
it more closely resembles that of the transition state. The stress
produced by such distortion of the substrate can further facilitate its
conversion to the transition state by weakening critical bonds.
Moreover, the transition state is stabilized by its tight binding to the
enzyme, thereby lowering the required energy of activation.
Factors that Affect Enzyme Activity
1. Concentration of Enzyme
 As the concentration of the enzyme is increased, the velocity of the
reaction proportionately increases. This property is used for
determining the activities of serum enzymes during the diagnosis of
diseases.
2. Concentration of Substrate
 In the presence of a given amount of enzyme, the rate of enzymatic

reaction increases as the substrate concentration increases until a
limiting rate is reached, after which further increase in the
substrate concentration produces no significant change in the
reaction rate. At this point, so much substrate is present that
essentially all of the enzyme active sites have substrate bound to
them.
 In other words, the enzyme molecules are saturated with substrate.

The excess substrate molecules cannot react until the substrate
already bound to the enzymes that have reacted and been released
(or been released without reacting).
3. Effect of Temperature
 The protein nature of the enzymes makes them extremely sensitive

to thermal changes. Enzyme activity occurs within a narrow range
of temperatures compared to ordinary chemical reactions. As you
have seen, each enzyme has a certain temperature at which it is
more active. This point is called the optimal temperature, which
ranges from 37 to 40°C.
 The enzyme activity gradually lowers as the temperature rises more

than the optimal temperature until it reaches a certain temperature
at which the enzyme activity stops completely due to the change of
its natural composition.
 On the other hand, if the temperature lowers below the optimal

temperature, the enzyme activity lowers until the enzyme reaches a
minimum temperature at which the enzyme activity is the least. The
enzyme activity stops completely at 0°C, but if the temperature
rises again, then the enzyme gets reactivated once more.
4. Effect of pH
 Each enzyme has a pH value that it works at with maximum
efficiency called the optimal pH. If the pH is lower or higher than
the optimal pH, the enzyme activity decreases until it stops
working. Most enzymes work at neutral pH 7.4.
5. Effect of Activators
 Some of the enzymes require certain inorganic metallic cations, like

Mg2+, Mn2+, Zn2+, Ca2+, Co2+, Cu2+, Na+, K+ etc., for their optimum
activity. Rarely, anions are also needed for enzyme activity, e.g. a
chloride ion (CI–) for amylase

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M8 – Properties of Protein

Physical Properties of Proteins

Proteins have large molecular weights and because of

The solubility of protein is influenced by pH. Solubility is lowest at an

Casein is the chief protein of milk. It is a phosphoprotein. It is

Gelatin is insoluble in cold water but soluble in hot water. It is a non-

If these changes occur to a small extent, denaturation can be

Factors that Cause Protein Denaturation:

1. Strong acids and bases

Changes in the pH can disrupt hydrogen bonds and salt bridges,

Alcohol coagulates all types of protein except prolamines. Alcohol

Changes in the salt concentration may result in the formation of a

Heat cleaves hydrogen bonds, so boiling a protein solution destroys

A fundamental task of proteins is to act as enzymes—catalysts that

Enzymes accelerate the rates of such reactions by well over a million-

Enzymes are characterized by two fundamental properties.

 First, they increase the rate of chemical reactions without themselves

The simplest model of enzyme-substrate interaction is the lock-and-

Factors that Affect Enzyme Activity

 In the presence of a given amount of enzyme, the rate of enzymatic

 In other words, the enzyme molecules are saturated with substrate.

 The protein nature of the enzymes makes them extremely sensitive

 The enzyme activity gradually lowers as the temperature rises more

 On the other hand, if the temperature lowers below the optimal

 Some of the enzymes require certain inorganic metallic cations, like

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