Download Protein Conformation and Function

Proteins
The Function of Proteins
Amino Acids
The Peptide Bond
Structure of Proteins
Myoglobin, Hemoglobin and Oxygen
Overview of Protein Structure and Function
Effect of Temperature and pH
1
• Proteins are among the “essential”
compounds necessary for the normal
functioning of a living system.
• The name is derived from the Greek word
“Proteios”, meaning first.
• All proteins are made from amino acids.
R O
NH2 C C
H OH
2
The function of proteins
Enzymes
Biological catalysts.
Antibodies
They fight off infection.
Transport
Move materials around
Ex. hemoglobin for O2.
Regulatory
As hormones, they control
metabolism.
Structural
coverings and support
skin, tendons, hair, nails, bone.
Movement
muscles, cilia, flagella.
3
Proteins are often gigantic in size
– Insulin:
Molecular Wt = 5700
– Hemoglobin:
Molecular Wt = 64,000
– Virus Proteins:
Molecular Wt = 40,000,000
4
Amino acids
All these proteins are made from the same
building blocks.
•
•
•
Twenty common amino acids.
All are -amino acids except proline.
A primary amine is attached to the  carbon.
-carbon - the carbon just after the acid.
H
|
R-C-COOH
|
NH2
5
Amino acids
Because an acid and base are both present, an
amino acid can form a +/- ion, called a
zwitterion.
H
|
R-C-COOH
|
NH2
H
|
R-C-COO
|
NH3+
How well it happens is based on pH and the
type of amino acid.
6
-Amino acids
Except for glycine, the  carbon is attached to
four different groups - it is a chiral center.
For Carbohydrates
We used the D- form.
For Amino Acids
We use the L- form.
COO+ |
H3N - C - H
|
R
The amino group is on the left side of the fischer
projection.
7
Classification of amino acids
• The -amino acid group is the same in each of
the amino acids.
• They are classified by the polarity of the side
chain (R).
Hydrophobic - water fearing
non-polar side chains
Hydrophilic - water loving
polar, neutral chains
negatively charged
positively charged
8
Neutral, nonpolar
side chains
glycine
H
|
H-C-COO|
+NH
3
leucine
H 3C
H
\
|
HC-CH2-C-COO/
|
+NH
H3C
3
H3C
valine
H
\ |
HC-C-COO/ |
H3C +NH3
alanine
H
|
CH3-C-COO|
+NH
3
9
Neutral, nonpolar
side chains
H 3C H
| |
H3C-CH2-CH-C-COO|
+NH
isoleucine
3
H
|
CH3 -S-CH2-CH2-C-COO|
+NH
3
methionine
H
|
-CH2-C-COO|
phenylalanine +NH3
proline
H 2C
|
H 2C
CH-COO|
+NH
2
H 2C
10
Polar, neutral amino acids
H
serine
|
HO-CH2-C-COO|
+NH
3
tyrosine
HO-
H
|
-CH2-C-COO|
+NH
3
HO H
|
|
CH3-CH-C-COO|
+NH
3
threonine
tryptophan
N
H
|
CH2-C-COO|
+NH
3
11
Polar, neutral amino acids
H
|
HS-CH2-C-COO|
+NH
3
cysteine
O
H
||
|
H2N-C-CH2-CH2-C-COO|
+NH
3
glutamine
O
H
||
|
H2N-C-CH2-C-COO|
+NH
3
asparagine
12
Acidic, polar side chains
Based on having a pH of 7.
glutamic acid
O
H
||
|
-O-C-CH -CH -C-COO2
2
|
+NH
3
aspartic acid
O
H
||
|
-O-C-CH -C-COO2
|
+NH
3
13
Basic, polar side chains
Based on a pH of 7.
+NH
H
|
|
H2N-C-N-CH2-CH2-CH2-C-COO|
+NH
arginine
3
2
H
|
+
H3N-CH2-CH2-CH2-CH2-C-COO|
+NH
lysine
3
H
|
CH2-C-COO|
+NH
N H
3
histidine
H
H N
+
14
Essential Amino Acids
• Proteins are constantly being produced in the
body for growth and repair.
• Of the 20 amino acids found in these proteins,
10 cannot be synthesized by the body.
• Arginine*
• Histidine *
• Isoleucine
• Leucine
• Lysine
Methionine
Phenylalanine
Threonine
Tryptophan
Valine
15
• Histidine is an essential amino acid for
infants, but apparently not for adults.
• Arginine is produced in the body but not
in sufficient quantities to meet protein
demand.
16
• Complete or Adequate Proteins: supply all of
the essential amino acids. (Animal Proteins)
• Incomplete Proteins: Low in one or more of the
essential amino acids (Vegetable Proteins)
Animal Proteins
Source
Type of Protein
Missing AA
Egg
Complete
None
Milk(Dairy)
Complete
None
Meat, fish
Complete
None
17
Source
Wheat
Corn
Vegetable Proteins
Type of Protein
Incomplete
Incomplete
Rice
Beans
Incomplete
Incomplete
Peas
Quinoa
Hemp
Incomplete
Complete
Complete
Missing AA
Lysine
Lysine &
Tryptophan
Lysine
Methionine
Tryptophan
Methionine
18
• A complete assortment of amino acids can be
obtained from a vegetable diet by pairing a
vegetable protein missing one essential amino
acid with a vegetable that contains it.
• The two vegetable proteins are called
complementary proteins.
– Ex. Rice and Beans
19
Amphoteric Properties of Amino
Acids
• Amphoteric substances act as acids or bases.
– They are acids when they donate protons.
– They are bases when they accept protons.
• Amino acids can act as acids or bases.
– When placed in an acidic solution (low pH),
they act as bases by accepting protons and
becoming positively charged.
– In basic solutions (high pH), they act as
acids by donating protons and becoming
negatively charged.
20
ALANINE
+
CH3
NH3 CH
O
C
-
O
H+
Acid
Solution
CH3
+
NH3 CH
CH3
O
NH2 CH
C
NET CHARGE
+1
Basic
Solution
OH-
O
C
OH
-
O
NET CHARGE
-1
21
• Amino Acids function as buffers because
they can neutralize small increases of
acid or base.
• Proteins are one of the major buffering
systems in the body.
22
ISOELECTRIC POINT (pI)
• A Zwitterion, which is electrically neutral
overall, can only exist at a specific pH value.
• This pH value, called the isoelectric point, is
different for each amino acid.
• Amino acids with hydrocarbon R groups attain
their isoelectric point between pH 5.0 and 7.0
• ex. Leucine pH = 6.0
• Basic amino acids need high pH values to
reach their isoelectric points.
• ex. Arginine pH = 10.8
23
• Acidic amino acids need low pH values.
• ex. Aspartic acid pH = 3.0
• Proteins also have isoelectric points
depending on the amino acids that make them
up.
– At their pH, proteins become insoluble in
water, clump together, and precipitate out of
solution.
24
The peptide bond
Proteins are polymers made up of amino acids.
Peptide bond - how the amino acids are
linked together to make a
protein.
H
|
H2NCCOOH +
|
R
H
|
H2NCCOOH
|
R’
H O
H
| ||
|
H2N - C - C - N - C - COOH
|
| |
R
H R’
This is a condensation reaction: H2O is eliminated.
25
• This bond between the two amino acids is
called a peptide bond.
• Two amino acids joined like this give what is
called a dipeptide.
CH3 O
NH2 CH C
Alanine
H
OH
+
O
NH CH C
H
Glycine
CH3 O
-H2O
H
O
NH2CH C NHCH C
OH
OH
alanylglycine (ala-gly)
These 2 amino acids could also link the other way.
26
• Any two amino acids can be joined in a similar
manner to form dipeptides.
• It doesn’t end here ! Each dipeptide still has a
COOH and an NH2 that can form new peptide
bonds.
• Adding a 3rd amino acid gives us a tripeptide.
• This process can be continued to get a
tetrapeptide, a pentapeptide, and so on until
we have a chain of hundreds or even
thousands of amino acids.
27
• The chains of amino acids are the proteins.
• The shorter chains are often called
polypeptides.
– Ex. Glucagon with 21 amino acids is a large
polypeptide.
– Insulin with 51 amino acids is a very small
protein.
• We will consider a protein to be a peptide
chain with a minimum of 30 amino acids.
28
Primary structure
of proteins
• Primary Structure: What are the amino acids
that make up the protein and how are they
arranged in the chain ? (The amino acid
sequence)
ala
gly
pro
arg
his
ser
asn
ile
thr
asp
leu
trp
cys
lys
tyr
gln
met
glu
phe
val
29
• The amino acids in a chain are often referred
to as residues.
– Ex. Ala-gly-lys 3 residue amino acids
• The amino acid residue with the free COOH
group is called the C-terminal, and the amino
acid residue with the free NH2 group is called
the N-terminal.
• Peptide and protein chains are always written
with the N-terminal residue on the left.
30
Peptides
N-terminal
residue
C-terminal
residue
H O
H O
H
| ||
| ||
|
H2N - C - C - NH - C - C - N - C - COOH
|
|
| |
R
R’
H R’’
peptide
linkages
31
• The continuing pattern of peptide linkages is
called the backbone of the protein molecule.
R
NHCH
O
C
R'
NHCH
O
R"
O
C
NHCH
C
The R groups are called the side chains.
The 20 different amino acid side chains
provide variety and determine the chemical
and physical properties.
32
• Each peptide and protein molecule in
biological organisms has a different sequence
of amino acids.
• It is this sequence that allows the protein to
carry out its function, whatever it might be.
• The number of different protein possibilities is
staggering.
– Ex. A tripeptide can have 20 different amino
acids at each position.
• 20 x 20 x 20 = 8000 possible tripeptides
33
• A typical protein with 60 amino acid residues
can have up to 2060 different arrangements.
• This means that there would be 1 x 1078
possibilities.
• 1,000,000,000,000,000,000,000,000,000,000,000,
000,000,000,000,000,000,000,000,000,000,000,0
00,000, 000,000.
34
Secondary structure
of proteins
Long chains of amino acids will commonly fold
or curl into a regular repeating structure.
Structure is a result of hydrogen bonding
between amino acids within the protein.
Common secondary structures are:
 - helix
 - pleated sheet
Secondary structure adds new properties to a
protein like strength, flexibility, ...
35
-Helix
One common type of
secondary structure.
Properties of -helix
include strength and
low solubility in water.
Originally proposed by
Pauling and Corey in
1951.
36
-Helix
C
||
O
H
|
N
C
||
OH
|
HN
|
N
H
|
N
C H
|| |
O N
H
|
N
C
||
O
C
||
O H
|
C
H
|| H N
C |
O |
|| N
C
O
N
||
C
O
||
O
Every amide hydrogen
and carbonyl oxygen is
involved in a hydrogen
bond.
There are 3.6 amino
acids in each turn.
Multiple strands may
entwine to make a
protofibril.
The R groups extend out from
the helical portion of the -helix
37
-Helix example
myosin head
myosin tail
ATP and actin
binding sites
thick filament
thin filament
actin
troponin
myosin/actin structure
Proteins used in muscle
38
-Pleated sheets
Another secondary structure for protein.
Held together by hydrogen bonding between
adjacent sheets of protein.
C
|
R
C
|
R
H
|
N
C
||
O
R
|
C
C
||
O
N
|
H
R
H |
| C
N
O
||
C
O
||
C
H
|
N
C
|
R
N
| C
H |
R
R
|
C
C
||
O
H
|
N
C
||
O
N
|
H
R
| O
C ||
C
O
||
C
C
|
R
N
| C
H |
R
39
-Pleated sheets
Silk fibroin - main protein of silk is an example
of a  pleated sheet structure.
Composed
primarily of
glycine and
alanine.
Stack like
corrugated
cardboard for
extra strength.
40
Linus Pauling
http://www.youtube.com/watch?v=yh9Cr5n21EE
41
Collagen
Family of related proteins.
About one third of all protein in humans.
Structural protein
Provides strength to bones, tendon, skin,
blood vessels.
Forms triple helix - tropocollagen.
42
•As an animal grows older,
the extent of cross-linking
increases and the meat
gets tougher.
•Treatment with boiling
water converts collagen to
gelatin. Therefore, cooking
meat converts part of the
tough connective tissue to
gelatin, making the meat
more tender. (ex. Stewing
chickens)
43
Tanning hides increases the degree of crosslinking, converting skin to leather.
44
• Wool, hair and muscle are all formed from
strands of alpha helixes.
• These proteins can be stretched because the
hydrogen bonds can be elongated and then
return to the original configuration.
• This is especially true for wool.
45
DISULFIDE BRIDGES
• Disulfide bridges are covalent bonds formed
when 2 cysteine units are oxidized to form a
cystine unit.
SH
SH
oxidation
reduction
S
S
The strength of this bond is much greater than that
of a hydrogen bond.
46
Fibrous proteins
• insoluble in water
• form used by connective tissues
• silk, collagen, -keratins
Globular proteins
• soluble in water
• form used by cell proteins
• 3-D structure - tertiary
47
Tertiary structure of proteins
• This refers to how the molecule is folded. It
makes the molecule very compact.
• Results from interaction of side chains.
• Protein folds into a tertiary structure.
• This is typical of proteins called globular.
– Found in egg and serum albumin,
hemoglobin and myoglobin, and enzymes
and antibodies.
48
Types of tertiary bonding
Possible side chain interactions:
- Similar solubilities
- Ionic attractions
- Attraction between + and - sidechains
- Covalent bonding
49
Tertiary structure
of proteins
Sulfide
Crosslink
Hydrophobic
interaction
-S-S-
-COO- H3N+-
Salt bridge
Hydrogen
bonding
Side chain interactions
Help maintain specific structure.
Oxidation of cysteine - crosslink formation.
O
||
HO-C-CH-CH2-SH
|
NH2
oxidation
[O]
O
||
HS-CH2-CH-C-OH
|
NH2
covalent
disulfide
bond
O
O
||
||
HO-C-CH-CH2-S - S-CH2-CH-C-OH +H2O
|
|
NH2
NH2
51
Hydrophobic attractions
Attractions between R groups of non-polar
amino acids.
Hydrogen bonding
Interaction between polar amino acid
R groups.
Ionic bonding
Bonding between oppositely charged amino
acid R groups.
52
CH 110 homework #13 & #14 due
Today!
CH 110 homework #15 & #16 due
Saturday, March 19th
Final Exam March 19th 9:00amnoon.
53
Quaternary structure
of proteins
Many proteins are not single peptide strands.
They are combinations of several proteins
- aggregate of smaller globular proteins.
Conjugated protein - incorporate another type
of group that performs a specific function.
- prosthetic group
54
Quaternary structure
of proteins
Aggregate structure
This example
shows four
different proteins
and two prosthetic
groups.
55
Hemoglobin and myoglobin
Hemoglobin
oxygen transport protein of red blood cells.
Myoglobin
oxygen storage protein of skeletal muscles.
Both proteins rely on the heme group as the
binding site for oxygen.
56
Myoglobin
Heme
57
Hemoglobin
2  chains
4 heme
2  chains
58
Hemoglobin and
oxygen transport
In the lungs, there is an abundance of O2 so
oxygen is picked up by the hemoglobin.
Hb + 4 O2
Hb(O2)4
When blood reaches the cells, there is a lack
of O2 so oxygen is given up by the
hemoglobin.
Hb + 4 O2
Hb(O2)4
59
Sickle cell anemia
Defective gene results in production of mutant
hemoglobin - one misplaced amino acid.
Glutamate is replaced by valine at 2 of the 547
positions.
Still transports oxygen but results in deformed
blood cells - elongated, sickle shaped.
Difficult to pass through capillaries.
Causes organ damage, reduced circulation.
Affects 0.4 % of American blacks.
60
Sickled cells
62
63
Comparison of normal and
sickle cell hemoglobin
Normal
Sickle
64
Summary of
protein structure
primary
secondary
H O
H O
H
| ||
| ||
|
H2N - C - C- NH - C - C - N - C - COOH
|
|
| |
R
R’
H R’’
tertiary
quaternary
65
Effect of temperature
and pH on proteins
• Both will alter the 3-D shape of a protein if you
go beyond a ‘normal’ range.
• Disorganized protein will no longer act as
intended - denatured. They become
biologically inactive.
• They will clump together - coagulate.
– Examples frying an egg
HCl in stomach
66
Denaturing of a protein
denatured
heat
or
acid
coagulated
heat
or
acid
67
•Changes in pH or
temperature may not
break any of the
peptide bonds.
•The primary structure
is maintained.
•If denaturation occurs
under extremely mild
conditions, the protein
may be restored to its
original shape.
68
COAGULATION:
•Most changes are so
drastic that the protein
remains denatured.
•Protein strands
become insoluble and
precipitate out of
solution (coagulate).
•Effects of coagulation
are irreversible.
69
Hydrolysis
• Will result in protein being reduced to simpler
peptides and amino acids.
• Amount of hydrolysis depends on pH, time and
temperature.
O
O
||
||
H2N - CH - C - NH - CH - C - OH + H2O
|
|
R
R’
H+ or
OH-
O
||
H2N - CH - C - OH
|
R
O
||
H2N - CH - C - OH
|
R’
70
• The effect of acid or base is to add or remove
H+ to ionic side groups.
– Salt bridges and hydrogen bonds are
disrupted.
• When a protein is placed in a strong acid or
base, coagulation may also occur.
– Ex. Cheese made from acid coagulated
protein (casein) which forms curds.
71
• Tannic acids in burn ointments cause protein
coagulation at the site of the burn.
• This forms a protective coating which acts as a
barrier to further loss of fluids.
• A household source of tannic acid is Tea.
– Applying dampened tea bags to a burn will
precipitate protein and form a protective
barrier over the wound.
72
Heat and UV Light
• Optimum temperature for most proteins is 37C.
• Very few proteins remain biologically active
above 50 C. (Some bacteria have protein that
remains stable up to 70 C and higher)
– Increased Thermal activity (heat or UV)
disrupts some of the hydrogen bonds and
attractions between non-polar side groups
that maintain secondary and tertiary
structures.
73
• When you cook food, you are denaturing
protein.
– Ex. Boiling an egg, frying a steak.
• High temperatures are used to disinfect
surgical instruments, gowns, and gloves.
– AUTOCLAVE.
74
ORGANIC SOLVENTS
• Solvents like Ethanol, isopropyl alcohol, and
acetone disrupt the hydrogen bonding of
proteins by forming their own hydrogen bonds
with the protein.
• These solvents are used as disinfectants.
– A 70% solution of ethanol or IPA can pass
thru cell walls of bacteria
– Once inside, they cause coagulation of the
bacterial proteins within the cell.
• A 90% solution isn’t nearly as effective. Why?
75
Heavy Metal Ions
• Metal ions like Ag+, Pb 2+, Hg 2+, etc.. cause
denaturation of protein.
• The heavy metals react with the disulfide
bonds and the carboxyl groups of acidic amino
acids.
• The denatured protein is insoluble and
precipitates out of solution.
– Ex. 1% AgNO3 placed in eyes of newborn to
kill the bacteria that causes gonorrhea.
76
Colloidal silver
77
Other Methods of Denaturation
• Agitation: Violent whipping action causes a
stretching of globular proteins which turns egg
whites into meringues and whipping creams
into toppings. How does cream of tartar work?
– Which would be best for beating eggs into
meringue: a glass bowl or a copper bowl?
– Why can canned pineapple be used in
gelatin deserts while fresh pineapple can’t
be used?
78
Protein gallery
Human growth hormone
596 residues
Originally obtained
from human cadavers.
It would cost $20,000
per year to treat one
child.
Now produced by
genetically engineered
bacteria.
79
Protein gallery
Immunoglobin FC - 262 residuals
A ‘Y” shaped protein actually composed of 4
protein chains linked by disulfide bonds.
antigen-binding
site
80
Protein gallery
Lipoprotein
116 residues
2 helical strands
81