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Amino Acids and Proteins
Amino Acids and Proteins
Amino Acids and Proteins
MEDICAL UNIVERSITY
BIOCHEMISTRY DEPARTMENT
ILLUSTRATED BIOCHEMISTRY
Tver, 2018
AMINO ACIDS
-amino acids -These are organic acids with at least a minimum of one of its
hydrogen atoms in the carbon chains substituted by an amino group.( Show the
radical, amino and carboxyl groups)
NH2
R |
C – H
|
COOH
Proteinogenous and Nonproteinogenous Amino acids
- Major proteinogenous (standard) amino acids. (Give the names of each amino
acid)
R R
1 H–
2 CH3 –
HO – CH2 –
(CH3)2 CH – 12
3 CH3 – CH –
(CH3)2 CH – CH2 – 13
4 |
CH3 – CH2 – CH –
5 | OH
CH3
14
6 HS – CH2 –
HOOC – CH2 –
7 HOOC – CH2 – CH2 – 15 CH3 – S – CH2 – CH2 –
NH2
|
- – C – H
|
COOH
8 16
NH2 – CO – CH2 –
9 NH2 – CO – CH2 – CH2 –
17
20
СООН
NH
-Glycine -Arginine
Alanine -Serine
-Valine -Threonine
Leucine -Cysteine
Isoleucine -Methionine
Aspartic acid -Phenylalanine
Glutamic acid -Tyrosine
Asparagine -Tryptophan
Glutamine -Histidine
Lysine -Proline
NH2
|
H2N – CH2 – CH – (CH2)2 – CH
| |
OH COOH
NH2 NH2
| |
H2N – (CH2)3 – CH HS – (CH2)2 – CH
| |
COOH COOH
NH2
|
NH2 – C – HN – (CH2)3 – CH
|| |
O COOH
Ornithine
Homocysteine
Citrulline
2
CLASSIFICATION OF PROTEINOGENOUS (STANDARD)
AMINO ACIDS
10
*The influence of pH on the ionization (charge) of an amino acid
-show how the charges of amino acids in basic and acidic media are changed.
-pH<7, the excess of hydrogen ions{H+}- acidic medium
-pH>7, the excess of hydroxyl ions {OH-}-basic medium
R
H+ | OH-
? (+) +
H3N – CH – COO– (+) ?
рН
7
COO- NH3+
| |
CH2 CH3 (CH2)4
| | |
H3 N – CH – COO–
+ H3+N – CH – COO– H3 N – СН – COO–
+
+ H+ ОH-
7
2,8 9,8
IEP IEP
11
•The amphoteric and buffer properties of amino acids
-What is amphoterity?
PROTEINS
They are ;
High –molecules
Nitrogen containing
Organic compounds (substances)
Made up of amino acids
joined in a chain
with the help of peptide bonds
and have a complex structural organization
(Explain all these characteristics)
-Elementary components of proteins (C50-54% , O21-23% , N15-17%, H6,5-7,3% , S0,5% )
-What are oligopeptides, polypeptides, proteins (up to 10 , 10-40 ,>40 amino acids) and
their molecular masses ( the average molecular mass of one (1) amino acid is 110 a.u ) ?
N.B a.u = atomic units
- Biological functions of proteins (Give examples of proteins with different functions and
give the characteristics of the actions of the proteins action in performing these functions)
12
- How can the different kinds of functions of proteins, their individual and
immune properties be explained?(Sequence and the number of the 20 proteinogenous
amino acids) .Explain your answer.
Primary ( Linear sequence of amino acids joined in a chain with the help of peptide
bonds)
R
|
H2N – CH – COOH
R1 R2 R3 Rn Rextreme
| | | |
H2N – CH – CОOH + H2N – CH – CОOH + H2N – CH – CОOH + ….….… + H2N – CH – C – OH
1 – Н2О 2 – Н2О 3 – Н2О n
R1 R2 R3 Rn O
| | | | //
H2N – CH – CО ––– NН – CH – CО ––– NН – CH – CО ………… NН – CH – C – OH
-Show :
13
-the radicals of the amino acids
-It is a way of folding of the polypeptide chain (Primary structure ) into a regulate -
spiral or a -structure ( kind of bond –Hydrogen bond)
–
О
||
– CO — NH– – C –– N –
|
Н +
β-structure
R4
-This is the folding of the -spiral (-helix) or the -structure in space (Globular,
Fibrillar proteins, explain their structure)
14
-Show how the tertiary structure is stabilized by ionic, hydrogen, covalent (disulfide)
bonds and hydrophobic interactions
Quaternary structure
-It is the joining the group of polypeptide chains with tertiary structures into a unit
functional protein molecule.
15
Which molecules contain information about the primary, secondary , tertiary and
quaternary structures of proteins and where are they located?
Explain the importance of the primary and quaternary structures of proteins in the
performance of their functional activities giving examples of ;
- denaturation of proteins
- sickle –cell anemia
- different structures and functions of myoglobin and hemoglobin
16
PHYSICAL AND CHEMICAL PROPERTIES OF PROTEINS
THE STRUCTURE, HYDRATE AND IONIC LAYERS OF PROTEIN
MOLECULES.
(0)
OR
Solubility of Proteins
Show and explain how the number (quantity) of amino acids , the hydrate layer and
the structure (conformation) affect the solubility of proteins
-neutral proteins
-acidic proteins.
Isoelectric Point of proteins
-using the diagram, show the value of the isoelectric point of the following proteins
(pH<7, >7, =7)
-basic proteins
-Explain how changes in the pH medium can affect the charge of a protein molecule
(<7, >7,=7)
1)
COO+ –
NH3
COO ––
2) COO+
NH3
COO+ –
3) NH3+
NH3
[H+] [OH –]
17
- Using given diagrams explain:
-the isoelectric state of a protein molecule
-the isoelectric point (pI) of protein molecules.
-What is a buffer?
-Which groups of atoms exhibit buffer properties?
-Explain the value of the buffer capacity of proteins.
Semipermeable
membrane
18
Salting-out and the denaturation of proteins
-Factors that stimulate the above mentioned processes (choose the factors for
denaturation and those for salting-out)
-High temperatures
-Vibration
-Salts of alkaline and alkaline-earth metals
-salts of heavy metals
-Mineral and organic acids
-Organic solvents
-Ionized radiation
-Explain the differences in the structure and function of protein molecules during
denaturation and salting-out.
CLASSIFICATION OF PROTEINS
Proteins are classified by their:
-electrochemical indications (acidic, basic and neutral).Explain and give
examples
-polarity ;polar (hydrophobic), nonpolar (hydrophilic), amphiphilic. Explain these
terms.
-function (transport, enzymes, hormones, antibodies etc)
19
-chemical composition(simple and complex)
Simple proteins(Describe the amino acid composition, the molecular mass, the
charge , the structure and functions of the following:)
-Protamines -Albumins
-Histones -Globulins
COMPLEX PROTEINS
Principle of classification (based on the name of the prosthetic group) and classes of
complex proteins.
2. Lipid-protein complexes
3. Carbohydro-protein
complexes(Glycoproteins and
Proteoglycans)
4. Phosphoproteins
5. Metalloproteins
6. Neucleoproteins
CHROMOPROTEINS
Cobamidproteins
Flavoproteins
20
-Prosthetic groups (heme, vitamin A, Vitamin B, FAD).
-Functions (participating in the transport of oxygen, in the processes of vision,
hemopoesis, redox reactions.)
HEMEPROTEINS
Classification of hemeproteins
- Myoglobin – Cytochromoxidase
- Hemoglobin -Catalase
– Peroxidase
Myoglobin
Hemoglobin (heme + globin)
4 (heme + globin) One protomer
Four protomers
13
Myoglobin
Globin is the protein component. Describe its molecular mass, the number of
amino acids, structure(secondary and tertiary), and the place of attachment of the
heme (Histidine)
globin
О2
Hemoglobin
-Similarities and differences between hemoglobin and myoglobin.(Compare in
according to the molecular mass, the structure and the affinity for oxygen.)
-Cooperativity during the binding of hemoglobin oxygen and the subsequent
changes in the affinity for oxygen.
Explain the biological importance of the different affinities of hemoglobin and
myoglobin for oxygen.
-Oxygen dissociation curves for hemoglobin and myoglobin in the lungs and other
tissues (in the skeletal muscles).
14
DERIVATIVES OF HEMOGLOBIN
- There are other groups of atoms that can attach to the heme of hemoglobin and
myoglobin, for examples CO, CO2, CN, OH. The complexes formed as a result are
named as follows:
- Oxyhemoglobin - Methhemoglobin
- Carboxyhemoglobin - Cianhemoglobin
15
-Carbhemoglobin.
CARBOHYDRO-PROTEIN COMPLEXES
Glycoproteins - The relation of the protein component to that of the prosthetic group
- 95:5.
16
- Carbohydrates of glycoproteins;
- Their chemical nature and structure.
17
CARBOHYDRATE COMPONENTS OF GLYCOPROTEINS
14
-Sulphur derivatives of glycosaminoglycans.
-Examples of proteoglycans, glycosaminoglycans and their biological functions in the
organism.
*Hyaluronic acid
*Chondrotin sulphate
*Dermatan sulphates
*Keratan sulphates
*Heparin
15
THE STRUCTURAL ORGANISATION OF THE INTERCELLULAR
MATRIX . (FRAGMENTS OF COMPLEXES OF HYALURONIC ACIDS
WITH PROTEOGLYCANS)
METALLOPROTEINS
*Transport
*Depot
*Structural functions
*Fermentative functions
These complexes are divided into two groups : free-lipid-protein complexes (blood
lipoproteins ) and structural lipoproteins (proteolipids of membranes).
Plasma lipoproteins
16
Pre-requisite for the formation of these complexes are NB; lipids are
hydrophobic
The structure of the lipoprotein complexes consists of hydrophobic and
hydrophilic components.
17
1 – according to the classification based on density .
2 - according to the classification based on electrophoretic mobility.
18
membrane of
the small
intestine
Density (g/ml) 0.92-0.98 0.96-1.00 1.00-1.06 1.06-1.21
Diameter of the More than 120 30-100 21-25 7-15
units (nm)
2% 3% 2% 10%
3%
18%
55%
7%
Chilo- VLDL
microns 10%
90%
3%
7%
22% 16%
4%
50%
42%
21% 27%
LDL HDL
8%
19
-Structural Lipoproteins (Proteolipids of cell membranes and their subunit
structures) NB ;These are membrane –bound lipoproteins.
-The protein –to-lipid relation in different membranes (the cell membrane, the
mitochondrial membrane, myelin-50:50, 80:20, 20:80.)
-The lipids of cell membranes (phospholipids, cholesterol, plasmalogen,
sphingolipids)
-The biological importance of proteins found in cell membranes (structural, antigens,
receptors, ferments, transport, etc)
-The unitary model of the cell membrane , ie, the “sandwich” model by Danielli and
Dawson (1931).The presence of a phospholipid bilayer , with pores covered by
different proteins.
-The liquid-mosaic model of the cell membrane by Singer and Nicholson (1972).The
phospholipid bilayer is of liquid consistency , and it has proteins both on the surface
and those that permeate (penetrate) the whole width of the bilayer.
Cholesterol decreases the fluidity of the membrane and helps in the structural
organization of the bilayer.
20
-Lipid and protein molecules of membranes are always in regular defined motions
(lateral displacement or turning)
-The asymmetry of membranes is due to the different structures, chemical
compositions and functions on the different sides of the membrane.
MECHANISMS USED FOR THE TRANSPORT OF LIPOPHILIC AND
HYDROPHILIC SUBSTANCES THROUGH THE CELL MEMBRANE
-Transport mechanisms according to the laws of diffusion.(Normal (I) and facilitated
(II)diffusion)
-The active transport of substances through their concentration gradient (protein-
carriers, ATP, symport (IIIa) and antiport (IIIb) .
The vesicular transport of macromolecules through the cell membrane (endo- (IV)
and exocytosis (V).)
21
22
THE BIOLOGICAL FUNCTIONS OF MEMBRANES (EXPLAIN).
23
NUCLEOPROTEINS
Deoxyribonucleoproteins (DNP) and Ribonucleoproteins (RNP).These are complex
proteins made up of simple proteins (protamines and histone)+DNA or RNA
Historical information
1868 - Meischer isolated and separated nucleic acids from the nucleus,
1943 - Avery, Macleod, MacCarthy found out that, the DNA of a virulent bacteria
converts a non-virulent culture of bacteria into virulent ones. DNA carries information
on heredity
1949 - Chargaff and his colleagues discovered the regularity (conformity) of the
formation of the DNA structure
1953-Watson and Crick created the DNA model
1967 - Kornberg synthesized the DNA of a virus
1970 - Corana synthesized an artificial(synthetic)gene
1978 - Arber, Smith, Nathanson discovered the phenomenon of DNA restriction
2000 - Scientists all over the world came close to totally decoding the human
DNA
-2,4-dioxypyrimidine (Uracil)
-5-methyluracil (Thymine )
-2-oxo-4-aminopyrimidine
-6-aminopurine (Adenine)
24
-2-amino-6-oxopurine (Guanine)
Ribonucleosides Deoxyribonucleosides
-Adenosine - deoxyadenosine
-Gyanosine - deoxyguanosine
-Cytidine - deoxycytidine
-Uridine - thymidine
Ribose-5-phosphate
-Principle of formation
25
-Thymidylic acid (thymine monophosphateTMP)
- Names of deoxyribonucleotides:
- Deoxyadenosinemonophosphate ( d-AMP etc)
- Unusual mononucleotides and their nitrogen bases (5-methyladenine, 1-
methyladenine).
The biological importance of these mononucleotides.
(Defense, regulatory)
NUCLEIC ACIDS
(These are polymers made up of different mononucleotides joined together by
phosphodiesteric bonds)
These acids may either contain ribose or deoxyribose and also they may either contain
Uracil or Thymine. Based on these nucleic acids may either be called RNA or DNA.
-Principle of formation –complex- esteric covalent bonds(3’,5’)between the ribose of
the previous mononucleotide (C3-OH) and the phosphoric acid of the next
mononucleotide.(C5-OH)
26
-linear sequence of mononucleotides in a polynucleotide chain.
-3’and 5’ terminals of nucleic acids
-Show and explain the given positions of the diagram on page 26
(V) The DNA does not change during the lifetime of an organism and does not
depend on nutrition or the external environment.
Results of the x-ray diffraction photograph and its analysis (the work by Maurice
Wilkins and Rosalind Franklin)
-Periodicity of the polymeric chain of the DNA along the 0,34 and 3,4nm axis
27
-Two polynucleidotide chain (double helical structure)
-Chains are antiparellel (5’,3’ and 3’,5’)
-External side of the spiral – phosphoric acid connected with a deoxyribose.
-Internal components of the spiral – nitrogen bases, which are located
complementary by each other (A::::::::::::T, G C)
28
The principle of formation:
-Histones + a fragment of the DNA (~200 pairs of mononuleotides ) =nucleosome
-During cell division, the genetic material is grouped in the form of chromosomes.
-During the active synthesis of proteins ,the chromosomes untwist and form active
chromatin
Deoxyribonucleic protein (DNP) =DNA + proteins
m-RNA (messenger-RNA)
-they are formed after the copying of information from the DNA.
-they serve as the template on which the specific amino acid sequence of a protein
molecule is built.
-they code one or more polypeptide chains
-the genetic code for m-RNA; triplet (codon)
-m-RNA + protein = informosome
r-RNA (ribosomal-RNA)
t-RNA (transfer-RNA)
29
-chemical composition
-quantity of mononucleotides
-the secondary structure of t-RNA (the formation of the “clover leaf” and the
functionally important parts)
1-the acceptor arm- CCA
2-the anticodon
3-pseudouridine arm (loop); this is the part used to attach to the ribosomes
(TC-loop)
4-dihydrouridine loop (arm) or the D-arm ,this is the point where the ferment
aa-t-RNA-synthetase attaches.This ferment is responsible for the joining of specific
amino acids to the given t-RNA. (D-loop).
-there are 20 proteinogenic (standard) amino acids and there are over 60 t-RNA.
(Explain the reason of the t-RNA excess).
*one amino acid is transported by 2-3 tRNA, but one t-RNA can only transport
a definite amino acid.
*aminoacyl-t-RNA (aa-t-RNA) is the active form in which amino acids are
transported.
30
PHYSICO-CHEMICAL PROPERTIES OF NUCLEIC ACIDS AND
NUCLEOPROTEINS
-Molecular mass
-Charges of the molecules
-Colloid properties
-Viscosity
-Ability to denaturate and renaturate
I II
renaturation
denaturation
GENETIC ENGINEERING
This is a direction of molecular genetics aimed at working out methods for the
receiving and transplanting of needed genes into the DNA of a host, thereby changing
the genetic properties of the host cell.
-Methods of genetic engineering (basic stages)
-receiving the gene of interesting DNA fragment
-the joining of this DNA to a vector molecule (a plasmid, etc)
-introducing the needed gene into the host-cells.
-selection of cells in which the reproduction of the introduced needed gene occurs.
31
-Cyclic mononucleotides(c-AMP, c-CMP) and their biological role (secondary
messengers)
32
TEMPLATE SYNTHESES (BIOSYNTHESIS OF DNA, RNA AND
PROTEINS)
Ways of transfer of genetic information:
Removal of RNA
primers and filling the
gap with DNA
polymerase
33
-What are the constituents of the newly formed DNA molecules?( the mother chain
and the newly formed daughter chain)
-Stages of replication
-Identifying the origin of replication and the unwinding of the DNA strands
(DNA-unwinding and DNA-binding proteins)
-Initiating the synthesis of DNA (primase, RNA-primer)
-Elongation (DNA-polymerase III, Okazaki fragments, DNA-polymerase I,
ligase)
-Winding (reconstitution) of the strands into a double helix structure (spiral).
-Termination (Reconstitution of chromatin)
*Predecessors of the daughter chains (RNA-primers, macroergs, Okazaki fragments)
*Leading (forward) and lagging (retrograde) strands. Explain the differences
between these two chains.
*DNA replication system (DNA polymerases, fragments, enzymes, etc)
*What is the speed of daughter chains formation?
*The frequency of mistakes during replication (10-10).
DNA REPARATION
34
1-Identifying the damaged part
2-Deletion of damaged part
3-Alignment of “needed” nucleotides
4-Rejoining (sewing together) of the repaired strand
-Genic Mutations
-Biological results of mutation :
Neutral
Negative (hereditary pathologies like sickle -cell anemia)
positive (factor of evolution)
Replication Transcription
Biosynthesis of DNA Biosynthesis of RNA
Thymine, deoxyribose Uracil, Ribose
The whole chromosome is copied Recopying of information from different
fragments of the chromosome(from the
genes)
35
A new daughter DNA copy is formed t-RNA, m-RNAand r-RNA are formed
Double strand molecule of daughter DNA Single –strand molecule of RNA
identical to the mother DNA
Stages of transcription
Splicing –Removal of the intervening sequences of introns from the transcript and
the splicing together of exons
36
-RNA polymerase
-RNA
-Viruses contain reverse transcriptases. The biological results of the functions of these
viral transcriptases are as follows: viral hepatitis, HIV, etc.
Structure of HIV
37
-Matrix of virus (p17,p18)
-Nucleocapsid (p24,p25)
-Contents of the nucleocapsid (RNA, reverse transcriptase, endonuclease)
-Genom of HIV-I (3structural and 5 regulatory genes)
-HIV binds with the glycoproteins on the external surface of the membrane of host -
cells
-Fusion of HIV membrane with the host cell .
The nucleocapsid of the virus then penetrates into the cytoplasm of the host cell.
-With the help of proteases, the viral RNA is released from the nucleocapsid
-The biosynthesis of the pro-viral DNA on the template of the viral RNA with the
help of reverse transcriptases
-Integration of the pro-viral DNA genom into the( DNA) genom of the host cell
-Latent period (there is no transcription of the pro-viral DNA). Diagnosis of HIV is
doubtful .
-Provocation (increase in body temperature, alcohol intoxication ,changes in
hormonal status) and the activation of transcription of the pro-viral DNA
A large number of viral RNA is formed .
-Production of all viral components with the help of the viral RNA and the
formation of daughter viruses
-Cytolysis of cell membranes which leads to the release of the viruses and the
infection of other cells of the host
-There is a rise of viral antigens in the blood and a corresponding rise in the level of
antibodies
-Clinical manifestations of the disease.
-Laboratory diagnosis of HIV infections
-Establishment of the fact of infection
-Defining the stages of pathology development.
-Establishing the prognosis of the disease and its effective treatment .
Infection is proved by the presence of the following substances in the blood serum:
-HIV antigens
-HIV antibodies
-Pro-viral DNA
- Methods of analysis
-Immunofermentative analysis
-Method of hybridization of nucleic acids with specific DNA probes
-Method of polymeric chain reactions
-Doubtful results of diagnosis and actions of the doctor (within3-6 months repeated
medical checks)
-Final diagnosis of HIV infection (clinics, epidemiological laboratory diagnosis )
BIOSYNTHESIS OF PROTEINS(TRANSLATION) AND ITS REGULATION
Stages of translation
I Activation of amino acids
38
II Initiation of translation
III Elongation
IV Termination
V Post translatory modification
- Participants of translation (ferments, amino acids ,tRNA, rRNA, mRNA, factors of
initiation, factors of elongation, factors of termination, macroergs, etc)
39
Components:
40
Ribosomes
m-RNA
f-met-t-RNA (N-formylmethionine)
Initiating factors (IF-1,IF-2,IF-3)
-GTP
-Initiating codons (AUG,GUG)
-Peptidyl part(P) of the ribosome
-Amino acyl (acceptor part ,A-part ) of the ribosome
-Chemical components of the initiating complex
III Elongation
Components:
-aa, tRNA, aa- tRNA………….
-Peptidyl transferase
-mRNA
-Ribosome
-GTP-translocase
-Elongation factors (EF-1,EF-1,,EF-2)
IV TERMINATION
41
-disintergration of the m-RNA
-the speed of the biosynthesis of proteins (polysomes-80 ribosomes)
COMPONENTS:
-structural genes
-regulatory genes
-promoter
-operator
-DNA-dependent RNA-polymerase
-m-RNA
-protein-repressor
42
-ferments
-substrates
-hormones
-end products
-The Jacob and Monod operon hypothesis is confirmed by incubating bacteria in a
medium containing either glucose() or amino acids() :
43
44
BIOCHEMISTRY OF THE IMMUNE SYSTEM
45
THE CLASSIFICATION OF IMMUNOGLOBULIS BASED ON THE TYPE OF
HEAVY CHAINS
IgA, IgG, IgD, IgM, IgE.
Classes of
antibodies IgA IgG IgM
(immunoglobulin,Ig)
The specificity of antibodies (this is defined by the structure of the variable portions)
VARIABILITY OF ANTIBODIES
Quaternary 2 light chains 2 heavy chains
structure
Structure of the
chains
VL IL CL VH IH CH1CH2
There are 2 domains in the There are 4 domains in the H-chain
L-chain
V-the variable part( they are various for different antibodies)C-the constant part. I- the
intermediate portion
Representation
of the antibody
46
Schematic representation of immunoglobins origin
Antibody
Surface Ig B-lympho-
cyte
MHC protein
B-lymphocyte B-lymphocyte
T-helper
B-lympho-
cyte
-In the human organism there are 107 different clones of B-lymphocytes. Each of
these cells has its own surface Ig.
-antigens that enter the body are bound by surface Ig’s of specific clones of B-
lymphocytes.
-the antigen together with the surface Ig and the MHC (Major Histocompatibility
Complex) protein move into the cell.(Formation of the endosome).
-The MHC of the B-lymphocyte changes conformation. (Processing)(3)
47
-The changed MHC protein moves to the surface of the B-lymphocyte and it is
recognized by T-Helpers. (4)
-T-Helpers secrete factors of proliferation.(4)
-These factors activate the proliferation specific clones of B-lymphocytes and
strengthens the synthesis of specific antibodies.
48
HUMORAL FACTORS OF THE IMMUNE SYSTEM
Complement- This is a family of specific blood-plasma proteins
-In normal physiological conditions ,the complement proteins are in the inactive state
in the blood plasma.
-There are two ways of activating them:
-the classical path; this form of activation arises as a result of the
binding of an antigen to an antibody
-the alternate path; this form of activation occurs in response to
polysaccharides or antigens.
-Both paths activate the C3 component of the complement system
-The cascade mechanism of activating the complement system is realized as a result
of the activities of proteases and the release of histamines
-The Functions of the Complement System
*Interaction with components of bacterial membranes
*Enhances the attachment of leucocytes and the phagocytosis of bacteria
*Lysis of bacteria-Components of the complement system penetrate into cells
bacterial membranes and make holes in it. The free movement of Na+ and Ca2+ ions
49
into, and K+ out of the cell ,causes the hydration and an increase in volume of the cell,
which leads to the destruction of the cell membrane.
50