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HBC1011 Biochemistry I
Trimester I, 2018/2019
Lecture 6 – Protein structure and
function
Ng Chong Han, PhD
MNAR1010, 06-2523751
chng@mmu.edu.my
Overview
• Protein function and functional group
• Amino acid: Chemical and Physical properties,
Types
• Polypeptide and peptide bond
• Primary structure
2
Protein function
• Protein – large biological molecules, consisting of one or more
long chains of amino acid residues.
• Functions
– catalysts
– transport and store other molecules such as oxygen
– provide mechanical support and immune protection
– generate movement
– transmit nerve impulse
– control growth and differentiation.
• Proteins differ in their sequence of amino acids, which is dictated
by the nucleotide sequence of their genes, and which usually
results in folding of the protein into a specific three-dimensional
structure that determines its activity.
3
The four levels of protein structure - 1
4
• Proteins are linear polymers built of monomer units called amino
acids, which are linked end to end.
• Primary structure - the sequence of linked amino acids
• Secondary structure - 3D structure formed by H bonds between
a.a. near one another
The four levels of protein structure - 2
5
• Tertiary structure - is formed by long-range interactions between a.a.
• Quaternary structure - the three-dimensional structure of a multi-
subunit protein and how the subunits fit together. Protein function
depends directly on this three-dimensional structure
Protein – Functional group
• Contains wide range of functional groups (on the side chains of
the a.a.)
e.g.: alcohol, thiol, thioether, carboxylic acids, carboxamides
• Chemically reactive
• Aid in the protein’s 3-D structure formation
• Contribute to the protein’s function
6
Protein properties
• Proteins can interact with one another and with other
biological macromolecules to form protein complex.
– Examples : DNA replication machinery
• Rigid: in cytoskeleton/in connective tissue
• Flexibility: hinges, spring, lever structures in macromolecular
assembly
7
Protein complex
Protein – 20 amino acids as building blocks
• Amino acids (a.a.) are amino derivatives of carboxylic acids, in
which amino (-NH2) and carboxyl (-COOH) functional groups,
along with a distinctive side-chain (R) specific to each amino acid
are bonded to the same carbon atom (α-carbon atom).
• The elements of an a.a. - carbon (C), hydrogen (H), oxygen (O),
and nitrogen (N), and other elements found in the side-chains (R).
8
α
side
chain
α-amino
group
α-carboxyl group
Non standard amino acids
• Pyrrolysine - found in methanogenic organisms and other
eukaryotes.
• Selenocysteine - present in many noneukaryotes as well as most
eukaryotes, not coded directly by DNA.
• N-formylmethionine (fMet) – is often the initial amino acid of
proteins in bacteria, mitochondria and chloroplast.
9
N-formylmethionine
Stereoisomerism
• 19 of the 20 a.a. - are chiral, or asymmetric molecule since 4
different groups bonded to it except for glycine whose R is H atom.
• Stereoisomers – compounds that have the same molecular formula
but differ in the arrangement or configuration
• A.a. may appear in the form of L (Latin laevus, left) or D (Latin
dexter, right) isomers, which comes from the ability of optically
active compounds to rotate polarized light to the left or the right.
.
10
D-Alanine and L-Alanine
are nonsuperimposable
mirror images.
Stereoisomerism
• Most a.a. are in L-amino acids. A simple mnemonic for correct L-
form is "CORN": when the Cα atom is viewed with the H in the
back, the residues read "CO-R-N" in a anti-clockwise direction.
11
Zwitterion
• Zwitterion is a neutral molecule with both positive and
negative electrical charges.
• Under the physiological pH range (6.8-7.4), a.a. are zwitterions, or
dipolar ions. The amino group is protonated (-NH3
+) because the
pKa is close to 9 while the carboxyl group is deprotonated/ionized
(-COO-) because the pKa is below 3.
12
NH2RCHCO2H NH3
+RCHCO2
−
un-ionized zwitterion
Ionization state of amino acid
• Ionization state of an a.a varies with pH
• In acid solution, a.a.’s amino group protonated (-NH3
+) & carboxyl group is not
dissociated (-COOH), predominantly positively charged.
• pH raised: carboxylic acid group give out a proton. (Zwitterionic form – both
charges)
• Until pH ~9: protonated amino group loses a proton , predominantly negatively
charged.
13
Predicting a.a. electrical
charges at a particular
pH is important since it
affects structure and
function.
pKa of amino acid
• pK1 = proton dissociation from the
carboxyl group
• pK2 = proton dissociation from the
amino group
• pKR (for some a.a. with R ionizable
group) = proton dissociation from the
R group
14
• Because a.a contain acidic & basic
functional group, they can be titrated.
Titration curve for
Alanine (R=CH3)
Ionic forms of alanine
Dissociation of the carboxyl group: At a low pH, both of these
groups are protonated. As the pH of the solution is raised, the –
COOH group of Form I can dissociate by donating a proton, H+ to
the medium. The release of a proton results in the formation of
the carboxylate group, –COO–. This structure is Form II, which is a
zwitterion, is the isoelectric form of alanine with a net charge of
zero.
15
H+ + H+ +
Ionic forms of alanine
Dissociation of the amino group: The second titratable group of
alanine is the amino (– NH3
+) group. Release of a proton from the
protonated amino group of Form II results in the fully
deprotonated form of alanine, Form III.
16
H+ + H+ +
17
Isoelectric point (pI)
• The isoelectric point (pI) - the pH at which a molecule carries no
net electrical charge.
• The net charge on the molecule is affected by pH and can
become more positively or negatively charged due to the gain or
loss, respectively, of protons (H+).
• The pI value can affect the solubility of a molecule at a given pH.
They have minimum solubility in water or salt solutions at the pH
that corresponds to their pI and often precipitate out of solution.
18
Isoelectric point (pI)
• At pH values between the two pKa values, the zwitterion
predominates, but coexists in equilibrium with small amounts of
net negative and net positive ions.
• At the exact midpoint between the two pKa values, the amount
of net negative and net positive ions balance, so that average net
charge of all forms present is zero.
• For amino acids such as alanine that possess monocarboxylic
acid and monoamino groups, pK1 and pK2 represent pKCOOH and
pKAmino respectively.
• For amino acids with charged side-chains, the pKa of the side-
chain is involved.
19
Isoelectric point (pI)
• For amino acids with
charged side-chains, the
pKa of the side-chain is
involved.
• The isoelectric points
reflect the nature of
ionizing R groups present.
20
Types of amino
acids
A.a. can be
categorized
based on the
properties of side
chains (R):
• Polar
(hydrophilic)
or Non-polar
(hydrophobic)
• Acidic or Basic
R group is
important for
protein function.
21
22
23
Non polar side chain (Hydrophobic) -1
24
• Aliphatic hydrocarbon group (the absence of a benzene ring)
• Glycine, Alanine, Branched-chain amino acids (Valine,
Leucine and Isoleucine)
Non polar side chain (Hydrophobic) - 2
25
• Aliphatic cyclic group
• Proline (more conformationally restricted than the other amino acids,
influence protein architecture)
• Aromatic side group
• Phenylalanine
• Tryptophan
• Sulfur-containing group
• Methionine (almost always the first amino acid in a protein)
Non polar side chain
26
• In proteins found in aqueous solutions––a
polar environment––the side chains of the
nonpolar amino acids tend to cluster
together in the interior of the protein.
• This phenomenon, the hydrophobic effect, is
the result of the hydrophobicity of the
nonpolar R-groups.
• The nonpolar R-groups fill up the interior of
the folded protein and help give it its 3D
shape.
• However, for proteins that are located in a
hydrophobic environment, such as a
membrane, the nonpolar R-groups are
found on the outside surface of the protein,
interacting with the lipid environment.
Proline – Structural support
27
• Proline differs from other amino acids in that proline’s side chain
and α-amino N form a rigid, five-membered ring structure.
• Proline has a secondary amino group. The unique geometry of
proline contributes to the formation of the fibrous structure of
collagen, and often interrupts the α-helices found in globular
proteins.
Polar side chain (Hydrophilic) - 1
Electrically neutral (uncharged) at neutral pH
• Hydroxyl group
– Serine and Threonine have –OH group attached to an aliphatic side chain
– Tyrosine has –OH group attached to an aromatic ring
• Thiol group
– Cysteine has –SH which can react with other thiol group to forms disulfide
(-S-S-) bridges in protein in an oxidation reaction. Important for protein
structure, metal ion binding, antioxidant
28
Disulfide bonds by oxidation of cysteine
29
• Reversible formation of a disulfide bond by the
oxidation of two molecules of cysteine.
• Disulfide bonds between Cys residues stabilize the
structures of many proteins, may act as redox sensor.
Side chains as sites of attachment for other
compounds
30
• The polar hydroxyl group of
serine, threonine, and, rarely,
tyrosine, can serve as a site of
attachment for structures such
as a phosphate group.
• In addition, the amide group of
asparagine, as well as the
hydroxyl group of serine or
threonine, can serve as a site of
attachment for oligosaccharide
chains in glycoproteins.
Polar side chain (Hydrophilic) - 2
• Amide group
– derived from carboxyl groups, in their side chains.
– Amide bonds do not usually ionize in the range of pH
encountered in biochemistry.
– Glutamine and asparagine can be considered derivatives of the
glutamic acid and aspartic acid, respectively.
31
Acidic side chain
• Aspartic acid and glutamic acid,
have carboxyl groups in their side
chains in addition to the one
present in all amino acids.
• A carboxyl group can lose a proton,
forming the corresponding
carboxylate anion, glutamate and
aspartate, respectively.
• They are negatively charged at
neutral pH.
32
Basic side chain
• Histidine, lysine and arginine
have basic side chains, and the
side chain is positively
charged at neutral pH.
• Lysine is capped by a primary
group and arginine by a
guanidinium group.
• Histidine contains an
imidazole group, which can
bind and release protons
during enzymatic reaction.
33
Essential amino acid
• Essential amino acid - an amino acid that cannot be
synthesized de novo (from scratch) by the organism
being considered, and therefore must be supplied in its
diet.
• If they are not taken through diet, they will not be
available for protein synthesis. In addition, some a.a. are
considered conditionally essential. Under certain
conditions like illness or stress the body might not be
able, or might be limited in the ability, to synthesize
them.
34
35
Polymerization of amino acids generates
protein
• Peptides are short chains of amino acid monomers linked by
covalent peptide (amide) bonds.
• The shortest peptides are dipeptides, consisting of 2 amino acids
joined by a single peptide bond, followed by tripeptides,
tetrapeptides, etc. A polypeptide is a long, continuous, and
unbranched peptide chain.
36
Constant
backbone
Variable side chain
Polymerization of amino acids generates
protein
37
(Condensation)
Polypeptides chain
• A polypeptide chain consists of :
• Main chain or backbone: regularly repeating part
• Distinctive side chain: variable part
• Backbone: rich in H-bonding potential
• Carbonyl group (C=O): good H-bond acceptor
• N-H group: good H-bond donor
38
Constant
backbone
Variable side chain
Peptide bonds formation
• Formed by linking the α-carboxyl group of one a.a to α-amino
group of another a.a via peptide bond/amide bond, causing the
release of H2O, hence it is a (dehydration) condensation reaction
and usually occurs between amino acids.
• The resulting C(O)NH bond is called a peptide bond, and the
resulting molecule is an amide. The four-atom functional group -
C(=O)NH- is called a peptide link.
39
Peptide bonds hydrolysis
• The equilibrium of reaction favor the side of hydrolysis rather
than synthesis
40
Reaction favor left side (reactant).
But hydrolysis rate is extremely slow.
• Synthesis of protein primary structure require input of free
energy
• Peptide bond: kinetically stable, lifetime of 1000 yrs in the
absence of catalyst
Primary structure
• Proteins – most natural polypeptide chains contain between
50 and 2000 amino acid residues.
• The mean molecular weight of an a.a. is 110 g mol-1, and so
the m.w. of most proteins are between 5500 and 220000 g
mol-1.
• The mass of protein – expressed in units of dalton, one
dalton is equal to one atomic mass unit.
– Eg a protein with a m.w. of 50,000 g mol-1 has a mass of 50,000 daltons
or 50 kd (kilodaltons)
41
42
• Each a.a. unit in a
polypeptide is called
residue.
• A polypeptide chain has
polarity because its ends
are different: an α-amino
group is present at one
end and an α-carboxyl
group at the other.
43
• The sequence of a.a. in
a polypeptide chain is
written starting with
the amino-terminal
(N-terminal) residue,
ending with the
carboxyl-terminal
(C-terminal residue)
Tyr-Thr-Gln-Trp-Ile is
different from
Ile-Trp-Gln-Thr-Tyr with
different chemical
properties.
Amino acid sequence
• In 1953, Frederick Sanger: a.a. seq. of insulin
• Landmark in biochemistry: protein has a precisely
defined a. a seq. consisting only the L-amino acid linked
by peptide bonds
• 2,000,000 protein seq. are known now
• Each protein has a unique, precisely defined a. a seq.
• Amino acid sequence a protein is referred to as primary
structure
44
Amino acid sequence of bovine insulin, sequenced by Frederick Sanger in 1959
Biochemist and molecular biologist
Dr. Frederick Sanger, two time
Nobel Prize winner.
The study of amino acid sequence
1. To elucidate the mechanism of action
– i.e. catalytic mechanism of an enzyme
2. To determine the 3-D structures of protein
– Uncovering the rules that govern the folding of
polypeptide chains
3. To study molecular pathology
– Severe/fatal disease resulted from a change in a
single a. a within the protein
4. To determine evolutionary history
– To trace molecular event in evolution from amino acid
sequence
46
Study questions
1. What are protein, peptide and polypeptides?
2. What is the difference between amine and amide?
3. What is a zwitterion?
4. Describe the classification of amino acid based on the side
chain properties.
5. Draw the basic structure of dipeptide with N- and C-terminal
end
6. How does peptide bond form?
7. What is isoelectric point?
47

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219101 lecture 6

  • 1. HBC1011 Biochemistry I Trimester I, 2018/2019 Lecture 6 – Protein structure and function Ng Chong Han, PhD MNAR1010, 06-2523751 chng@mmu.edu.my
  • 2. Overview • Protein function and functional group • Amino acid: Chemical and Physical properties, Types • Polypeptide and peptide bond • Primary structure 2
  • 3. Protein function • Protein – large biological molecules, consisting of one or more long chains of amino acid residues. • Functions – catalysts – transport and store other molecules such as oxygen – provide mechanical support and immune protection – generate movement – transmit nerve impulse – control growth and differentiation. • Proteins differ in their sequence of amino acids, which is dictated by the nucleotide sequence of their genes, and which usually results in folding of the protein into a specific three-dimensional structure that determines its activity. 3
  • 4. The four levels of protein structure - 1 4 • Proteins are linear polymers built of monomer units called amino acids, which are linked end to end. • Primary structure - the sequence of linked amino acids • Secondary structure - 3D structure formed by H bonds between a.a. near one another
  • 5. The four levels of protein structure - 2 5 • Tertiary structure - is formed by long-range interactions between a.a. • Quaternary structure - the three-dimensional structure of a multi- subunit protein and how the subunits fit together. Protein function depends directly on this three-dimensional structure
  • 6. Protein – Functional group • Contains wide range of functional groups (on the side chains of the a.a.) e.g.: alcohol, thiol, thioether, carboxylic acids, carboxamides • Chemically reactive • Aid in the protein’s 3-D structure formation • Contribute to the protein’s function 6
  • 7. Protein properties • Proteins can interact with one another and with other biological macromolecules to form protein complex. – Examples : DNA replication machinery • Rigid: in cytoskeleton/in connective tissue • Flexibility: hinges, spring, lever structures in macromolecular assembly 7 Protein complex
  • 8. Protein – 20 amino acids as building blocks • Amino acids (a.a.) are amino derivatives of carboxylic acids, in which amino (-NH2) and carboxyl (-COOH) functional groups, along with a distinctive side-chain (R) specific to each amino acid are bonded to the same carbon atom (α-carbon atom). • The elements of an a.a. - carbon (C), hydrogen (H), oxygen (O), and nitrogen (N), and other elements found in the side-chains (R). 8 α side chain α-amino group α-carboxyl group
  • 9. Non standard amino acids • Pyrrolysine - found in methanogenic organisms and other eukaryotes. • Selenocysteine - present in many noneukaryotes as well as most eukaryotes, not coded directly by DNA. • N-formylmethionine (fMet) – is often the initial amino acid of proteins in bacteria, mitochondria and chloroplast. 9 N-formylmethionine
  • 10. Stereoisomerism • 19 of the 20 a.a. - are chiral, or asymmetric molecule since 4 different groups bonded to it except for glycine whose R is H atom. • Stereoisomers – compounds that have the same molecular formula but differ in the arrangement or configuration • A.a. may appear in the form of L (Latin laevus, left) or D (Latin dexter, right) isomers, which comes from the ability of optically active compounds to rotate polarized light to the left or the right. . 10 D-Alanine and L-Alanine are nonsuperimposable mirror images.
  • 11. Stereoisomerism • Most a.a. are in L-amino acids. A simple mnemonic for correct L- form is "CORN": when the Cα atom is viewed with the H in the back, the residues read "CO-R-N" in a anti-clockwise direction. 11
  • 12. Zwitterion • Zwitterion is a neutral molecule with both positive and negative electrical charges. • Under the physiological pH range (6.8-7.4), a.a. are zwitterions, or dipolar ions. The amino group is protonated (-NH3 +) because the pKa is close to 9 while the carboxyl group is deprotonated/ionized (-COO-) because the pKa is below 3. 12 NH2RCHCO2H NH3 +RCHCO2 − un-ionized zwitterion
  • 13. Ionization state of amino acid • Ionization state of an a.a varies with pH • In acid solution, a.a.’s amino group protonated (-NH3 +) & carboxyl group is not dissociated (-COOH), predominantly positively charged. • pH raised: carboxylic acid group give out a proton. (Zwitterionic form – both charges) • Until pH ~9: protonated amino group loses a proton , predominantly negatively charged. 13 Predicting a.a. electrical charges at a particular pH is important since it affects structure and function.
  • 14. pKa of amino acid • pK1 = proton dissociation from the carboxyl group • pK2 = proton dissociation from the amino group • pKR (for some a.a. with R ionizable group) = proton dissociation from the R group 14 • Because a.a contain acidic & basic functional group, they can be titrated. Titration curve for Alanine (R=CH3)
  • 15. Ionic forms of alanine Dissociation of the carboxyl group: At a low pH, both of these groups are protonated. As the pH of the solution is raised, the – COOH group of Form I can dissociate by donating a proton, H+ to the medium. The release of a proton results in the formation of the carboxylate group, –COO–. This structure is Form II, which is a zwitterion, is the isoelectric form of alanine with a net charge of zero. 15 H+ + H+ +
  • 16. Ionic forms of alanine Dissociation of the amino group: The second titratable group of alanine is the amino (– NH3 +) group. Release of a proton from the protonated amino group of Form II results in the fully deprotonated form of alanine, Form III. 16 H+ + H+ +
  • 17. 17
  • 18. Isoelectric point (pI) • The isoelectric point (pI) - the pH at which a molecule carries no net electrical charge. • The net charge on the molecule is affected by pH and can become more positively or negatively charged due to the gain or loss, respectively, of protons (H+). • The pI value can affect the solubility of a molecule at a given pH. They have minimum solubility in water or salt solutions at the pH that corresponds to their pI and often precipitate out of solution. 18
  • 19. Isoelectric point (pI) • At pH values between the two pKa values, the zwitterion predominates, but coexists in equilibrium with small amounts of net negative and net positive ions. • At the exact midpoint between the two pKa values, the amount of net negative and net positive ions balance, so that average net charge of all forms present is zero. • For amino acids such as alanine that possess monocarboxylic acid and monoamino groups, pK1 and pK2 represent pKCOOH and pKAmino respectively. • For amino acids with charged side-chains, the pKa of the side- chain is involved. 19
  • 20. Isoelectric point (pI) • For amino acids with charged side-chains, the pKa of the side-chain is involved. • The isoelectric points reflect the nature of ionizing R groups present. 20
  • 21. Types of amino acids A.a. can be categorized based on the properties of side chains (R): • Polar (hydrophilic) or Non-polar (hydrophobic) • Acidic or Basic R group is important for protein function. 21
  • 22. 22
  • 23. 23
  • 24. Non polar side chain (Hydrophobic) -1 24 • Aliphatic hydrocarbon group (the absence of a benzene ring) • Glycine, Alanine, Branched-chain amino acids (Valine, Leucine and Isoleucine)
  • 25. Non polar side chain (Hydrophobic) - 2 25 • Aliphatic cyclic group • Proline (more conformationally restricted than the other amino acids, influence protein architecture) • Aromatic side group • Phenylalanine • Tryptophan • Sulfur-containing group • Methionine (almost always the first amino acid in a protein)
  • 26. Non polar side chain 26 • In proteins found in aqueous solutions––a polar environment––the side chains of the nonpolar amino acids tend to cluster together in the interior of the protein. • This phenomenon, the hydrophobic effect, is the result of the hydrophobicity of the nonpolar R-groups. • The nonpolar R-groups fill up the interior of the folded protein and help give it its 3D shape. • However, for proteins that are located in a hydrophobic environment, such as a membrane, the nonpolar R-groups are found on the outside surface of the protein, interacting with the lipid environment.
  • 27. Proline – Structural support 27 • Proline differs from other amino acids in that proline’s side chain and α-amino N form a rigid, five-membered ring structure. • Proline has a secondary amino group. The unique geometry of proline contributes to the formation of the fibrous structure of collagen, and often interrupts the α-helices found in globular proteins.
  • 28. Polar side chain (Hydrophilic) - 1 Electrically neutral (uncharged) at neutral pH • Hydroxyl group – Serine and Threonine have –OH group attached to an aliphatic side chain – Tyrosine has –OH group attached to an aromatic ring • Thiol group – Cysteine has –SH which can react with other thiol group to forms disulfide (-S-S-) bridges in protein in an oxidation reaction. Important for protein structure, metal ion binding, antioxidant 28
  • 29. Disulfide bonds by oxidation of cysteine 29 • Reversible formation of a disulfide bond by the oxidation of two molecules of cysteine. • Disulfide bonds between Cys residues stabilize the structures of many proteins, may act as redox sensor.
  • 30. Side chains as sites of attachment for other compounds 30 • The polar hydroxyl group of serine, threonine, and, rarely, tyrosine, can serve as a site of attachment for structures such as a phosphate group. • In addition, the amide group of asparagine, as well as the hydroxyl group of serine or threonine, can serve as a site of attachment for oligosaccharide chains in glycoproteins.
  • 31. Polar side chain (Hydrophilic) - 2 • Amide group – derived from carboxyl groups, in their side chains. – Amide bonds do not usually ionize in the range of pH encountered in biochemistry. – Glutamine and asparagine can be considered derivatives of the glutamic acid and aspartic acid, respectively. 31
  • 32. Acidic side chain • Aspartic acid and glutamic acid, have carboxyl groups in their side chains in addition to the one present in all amino acids. • A carboxyl group can lose a proton, forming the corresponding carboxylate anion, glutamate and aspartate, respectively. • They are negatively charged at neutral pH. 32
  • 33. Basic side chain • Histidine, lysine and arginine have basic side chains, and the side chain is positively charged at neutral pH. • Lysine is capped by a primary group and arginine by a guanidinium group. • Histidine contains an imidazole group, which can bind and release protons during enzymatic reaction. 33
  • 34. Essential amino acid • Essential amino acid - an amino acid that cannot be synthesized de novo (from scratch) by the organism being considered, and therefore must be supplied in its diet. • If they are not taken through diet, they will not be available for protein synthesis. In addition, some a.a. are considered conditionally essential. Under certain conditions like illness or stress the body might not be able, or might be limited in the ability, to synthesize them. 34
  • 35. 35
  • 36. Polymerization of amino acids generates protein • Peptides are short chains of amino acid monomers linked by covalent peptide (amide) bonds. • The shortest peptides are dipeptides, consisting of 2 amino acids joined by a single peptide bond, followed by tripeptides, tetrapeptides, etc. A polypeptide is a long, continuous, and unbranched peptide chain. 36 Constant backbone Variable side chain
  • 37. Polymerization of amino acids generates protein 37 (Condensation)
  • 38. Polypeptides chain • A polypeptide chain consists of : • Main chain or backbone: regularly repeating part • Distinctive side chain: variable part • Backbone: rich in H-bonding potential • Carbonyl group (C=O): good H-bond acceptor • N-H group: good H-bond donor 38 Constant backbone Variable side chain
  • 39. Peptide bonds formation • Formed by linking the α-carboxyl group of one a.a to α-amino group of another a.a via peptide bond/amide bond, causing the release of H2O, hence it is a (dehydration) condensation reaction and usually occurs between amino acids. • The resulting C(O)NH bond is called a peptide bond, and the resulting molecule is an amide. The four-atom functional group - C(=O)NH- is called a peptide link. 39
  • 40. Peptide bonds hydrolysis • The equilibrium of reaction favor the side of hydrolysis rather than synthesis 40 Reaction favor left side (reactant). But hydrolysis rate is extremely slow. • Synthesis of protein primary structure require input of free energy • Peptide bond: kinetically stable, lifetime of 1000 yrs in the absence of catalyst
  • 41. Primary structure • Proteins – most natural polypeptide chains contain between 50 and 2000 amino acid residues. • The mean molecular weight of an a.a. is 110 g mol-1, and so the m.w. of most proteins are between 5500 and 220000 g mol-1. • The mass of protein – expressed in units of dalton, one dalton is equal to one atomic mass unit. – Eg a protein with a m.w. of 50,000 g mol-1 has a mass of 50,000 daltons or 50 kd (kilodaltons) 41
  • 42. 42 • Each a.a. unit in a polypeptide is called residue. • A polypeptide chain has polarity because its ends are different: an α-amino group is present at one end and an α-carboxyl group at the other.
  • 43. 43 • The sequence of a.a. in a polypeptide chain is written starting with the amino-terminal (N-terminal) residue, ending with the carboxyl-terminal (C-terminal residue) Tyr-Thr-Gln-Trp-Ile is different from Ile-Trp-Gln-Thr-Tyr with different chemical properties.
  • 44. Amino acid sequence • In 1953, Frederick Sanger: a.a. seq. of insulin • Landmark in biochemistry: protein has a precisely defined a. a seq. consisting only the L-amino acid linked by peptide bonds • 2,000,000 protein seq. are known now • Each protein has a unique, precisely defined a. a seq. • Amino acid sequence a protein is referred to as primary structure 44
  • 45. Amino acid sequence of bovine insulin, sequenced by Frederick Sanger in 1959 Biochemist and molecular biologist Dr. Frederick Sanger, two time Nobel Prize winner.
  • 46. The study of amino acid sequence 1. To elucidate the mechanism of action – i.e. catalytic mechanism of an enzyme 2. To determine the 3-D structures of protein – Uncovering the rules that govern the folding of polypeptide chains 3. To study molecular pathology – Severe/fatal disease resulted from a change in a single a. a within the protein 4. To determine evolutionary history – To trace molecular event in evolution from amino acid sequence 46
  • 47. Study questions 1. What are protein, peptide and polypeptides? 2. What is the difference between amine and amide? 3. What is a zwitterion? 4. Describe the classification of amino acid based on the side chain properties. 5. Draw the basic structure of dipeptide with N- and C-terminal end 6. How does peptide bond form? 7. What is isoelectric point? 47