Transcript Protein: Amino Acids - University of Brawijaya

Protein: Amino Acids
Aulanni’am
Biochemistry Laboratory
Chemistry Departement
Brawijaya University
Aulani "Biokimia" Presentation 5
•
Proteins have an amino group, an acid, a
hydrogen, carbon molecule and a carbon
side chain.



Protein means primary or first and are necessary for
life.
Amino means contains nitrogen (NH2).
Proteins can also contain sulfur, phosphorus or iron.
Aulani "Biokimia" Presentation 5
Amino Acids
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L-Form Amino Acid Structure
COO
Carboxylic group
Amino group
+
H 3N
a
R group
H
H = Glycine
CH3 = Alanine
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Proton： abundant and small, affects the charge of a molecule
lone pair
electrons
Amino
High pKa Low
N H
H+
H+
N H
H
H
Low pKa High
Carboxylic
C
O H
O
O
C
O
H+
Ampholyte contains both positive and negative groups on its molecule
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Acidic environment Neutral environment
Alkaline environment
pK2 ~ 9
NH2 H+
R-C-H
COOH
NH2 H+
R-C-H
COOpK1 ~ 2
+1
NH2
R-C-H
COO-
5.5
0
Isoelectric point
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-1
Amino Acids Have Buffering Effect
pH 12
★
pK2
Isoelectric point =
pI
9
NH2 H+
6
H-C-R
COO-
3
★
pK1 + pK2
2
0
[OH] →
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pK1
Environment pH vs Protein Charge
Buffer pH
10
9
8
7
Isoelectric point,
pI
+
6
5
4
3
0
-
Net Charge of a Protein
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pKa of Amino Acid Residues
Residues on amino acids can release or accept protons
a -COOH
R -COOH
His -Imidazole·H+
Cys -SH
Tyr -OH
a -NH3+
R -NH3+
a -COOR -COO-
+ H+ pKa = 1.8~2.4
+ H+ pKa = 3.9~4.3
His -Imidazole + H+ pKa = 6.0
+ H+ pKa = 8.3
Cys -S+ H+ pKa = 10
Tyr -Oa -NH2
R -NH2
+ H+ pKa = 8.8~11
+ H+ pKa = 10~12.5
Smaller pKa releases proton easier
Only His has the residue with a neutral pKa (imidazole)
pKa of a carboxylic or amino groups is lower than pKa of the R residues
Aulani "Biokimia" Presentation 5
pKa of Amino Acids
Amino acids
Gly
G
Ala
A
Val
V
Leu
L
Ile
I
Ser
S
Thr
T
Met M
Phe
F
Trp
W
Asn
N
Gln
Q
Pro
P
Asp
D
Glu
E
His
H
Cys
C
Tyr
Y
Lys
K
Arg
R
-COOH -NH2
2.34
2.34
2.32
2.36
2.36
2.21
2.63
2.28
1.83
2.38
2.02
2.17
1.99
2.09
2.19
1.82
1.71
2.20
2.18
2.17
9.60
9.69
9.62
9.68
9.68
9.15
10.4
9.21
9.13
9.39
8.80
9.13
10.6
9.82
9.67
9.17
10.8
9.11
8.95
9.04
-R
pH
two pKa
pK2
pI
pK1
pK1 + pK2
2
three pKa
3.86
4.25
6.0
8.33
10.07
10.53
12.48
pK3
pK2
pK1
?
?
pI ?
[OH-]
Aulani "Biokimia" Presentation 5
H
first
HOOC-CH2-C-COOH
NH3+
+1
pK1 = 2.1
H
second
HOOC-CH2-C-COO-
Aspartic acid
Isoelectric point is the average
of the two pKa flanking the
zero net-charged form
2.1 + 3.9
= 3.0
2
0
NH3+
Isoelectric point
pK2 = 3.9
H
-OOC-CH -C-COO2
-1
NH3+ third
pK3 = 9.8
H
-OOC-CH -C-COO2
NH2
-2
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-2
pK3
-1
pK2
pK1
0
+1
[OH]
Amino Acids


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Nonessential amino acids
 a.k.a dispensable amino acids can be made within
the body
Essential amino acids
 a.k.a indispensable amino acids must be obtained
from foods
Conditionally essential amino acids are needed from
food sources if the building blocks to make them are
not available.
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An Essential Amino Acid
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Nonessential Amino Acids
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
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Most amino acids are neutral with an aliphathic (single
chain) or aromatic chain.
Two are dibasic with two amino groups:
 Histidine and arginine
A few are diacidic and are commonly used as
components of proteins in cell membranes
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Proteins


Peptide bonds connect the acid end of one
amino acid with the amino end of another.
They are the links that form a protein chain,
which can be simple or very complex.
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Dipeptide
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Formation of Peptide Bonds by Dehydration
Amino acids are connected head to tail
NH2
1
COOH
NH2
2
COOH
Dehydration
Carbodiimide
-H2O
O
NH2
1
C N
2
H
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COOH


A Tripeptide
consists of three amino acids
linked together.
When there are three or more amino acids, the
protein starts to form three dimensional shapes.
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Protein structure

Remember Starch?:


glucose+glucose+glucose+glucose+glucose…
Meet Protein:
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amino acid+amino acid+amino acid+amino acid…
Made of 20 different amino acids bonded together
in different sequences to form many SPECIFIC
proteins.
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Amino acids

Essential (10)
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Phenylalanine
Valine
Threonine
Tryptophan
Isoleucine
Methionine
Histidine
Arginine
Leucine
Lysine

Nonessential (10)
 Alanine
 Asparagine
 Aspartic acid
 Cysteine
 Glutaminc acid
 Gluatmine
 Glycine
 Proline
 Serine
 Tyrosine
Conditionally essential (3)
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
Cysteine
Glutamine
Tyrosine
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Elemental composition of protein
Element
%
Carbon
Hydrogen
Nitrogen
Oxygen
Sulfur
Phosphorous
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51.0 – 55.0
6.5 – 7.3
15.5 – 18.0
21.5 – 23.5
0.5 – 2.0
0.0 – 1.5
Classification of amino acids


Essential amino acid
 One that the body is unable to make or can only
make in inadequate quantities
 Need to be consumed from the diet
 8-10 essential amino acids
Nonessential amino acid
 One that the body can make in large enough
quantities
 Made from essential amino acids
 Not necessary to consume these in the diet
 10-12 nonessential amino acids
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Classification, cont.

Conditionally essential amino acid.
One that can become essential in certain
physiologic conditions
 3 of these
Example: Tyrosine becomes essential in people with
“Phenylketonuria (PKU)”


Aulani "Biokimia" Presentation 5
Structure levels in proteins
primary structure
amino acid sequence
secondary structure
spatial arrangement of amino acids within a polypeptide chain (a-helix, ß-sheet, unstructured turns)
tertiary structure
spatial arrangement of secondary structural elements
quatery structure
subunit interactions
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Amino acids are constituents of
proteins
there are about 20 proteinogenic amino acids
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Structure of amino acids
Central
carbon

H
H
O
H N
C
C OH
H
Amino
group
Side
chain
(R)
Different side chains
make different amino
acids
Acid
group
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Likage of amino acids
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Primary structure of a protein

It is the sequence of amino acids that makes each
protein different from the next
Peptide Bonds
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Dipeptide = 2 amino acids
Tripeptide = 3 amino acids
Polypeptide = many amino acids
Most proteins have many 100 amino acids
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Secondary structures
a-helix
ß-sheet
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Secondary structure
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Alignment of polypeptides as a right-hand alpha helix
Stabilized by hydrogen bonds between carboxyl (C=O)
and imido (NH) groups
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Unstructured turns connect
secondary structural elements
pronounced turn
limited turn
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Tertiary structure
helices are shown in yellow,
carbon backbone is shown in
black
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Tertiary structure


Three dimensional folding and coiling of
polypeptide into globular 3-D structure
Caused by additional chemical
interactions among side chains

Disulfide bonds
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Quaternary structure
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Interactive folding of
several polypeptide chains
together to form a
“single” functional protein
Functional proteins also
might
incorporate
minerals
or
other
nonprotein components
Final
shape
and
components
determine
function of protein
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Quartery structure
T-form
compact structure
R-form
relaxed structure
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
Polypeptide
Copyright 2005 Wadsworth Group, a division of Thomson Learning
Aulani "Biokimia" Presentation 5
Proteins
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Amino acid sequences can vary resulting in almost an
endless number of combinations.
Each protein’s sequence is determined by the DNA
As each amino acid has unique chemical
characteristics and electrical charges, the resulting
shapes can be very complex.
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Protein shape
and function
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Enzymes

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Proteins that catalyze (speed up) chemical reactions
without being used up or destroyed in the process.
Anabolic (putting things together) and catabolic
(breaking things down) functions.
Examples
 Digestion
 Salivary amylase
 Trypsin
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Hormones

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
Chemical messengers that
are made in one part of the
body but act on cells in
other parts of the body.
Note that "steroid
hormones" are not proteins!
Examples
 Insulin
 CCK
 Some reproductive
hormones
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Immune function (antibodies)

Antibodies are proteins that attack and
inactivate bacteria and viruses that cause
infection.
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Denaturation
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Protein denaturation happens when a protein changes
its shape, usually uncoiling.
This changes its function and properties.
An egg is mostly liquid until cooked. Milk becomes
yogurt or or cheese when acids or enzymes are added.
Heat, acids, bases, alcohol, heavy metals, enzymes or
other agents can cause denaturation.
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Protein Digestion
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
Stomach releases HCL, which denatures (uncoils)
protein strands and converts the inactive form of
pepsinogen into the active form pepsin.
Pepsin breaks the proteins into smaller polypeptides
Pepsin is one of thousands of enzymes, which allows
chemical reactions to take place in the body without
being affected itself.
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Small Intestine
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
Releases pancreatic and intestinal proteases.
These hydolyze the polypeptides further into
tripeptides, dipeptides and finally amino
acids, which are actively transported into SI
cells and then released into the blood stream.
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Protein Digestion in
the GI Tract
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Protein Absorption
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Carriers - cells of the villi of the SI have gates
through which carrier substances transport the amino
acids.
Capillaries, which are the smallest branches of the
circulatory system carry the free amino acids from
the villi throughout the body.
Absorption misconceptions
 Enzyme/amino acid supplements
Aulani "Biokimia" Presentation 5
• Messenger
RNA
from
the
nucleus
and Ribosomes within cells assemble the free amino
acids into proteins


As the ribosome moves along the mRNA, an
enzyme bonds one amino acid to another.
40 to 100 amino acids can be added to a
growing protein strand in one second.
Aulani "Biokimia" Presentation 5
Protein Synthesis
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DNA in the cell nucleus gives mRNA the instructions.
mRNA goes into the cellular fluid and attaches itself
to ribosomes
transfer RNA carries free amino acids to the mRNA
Ribosomes move along the mRNA allowing enzymes to
bond one amino acid to another until the completed
protein is finished and released.
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Protein Synthesis
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
Sequencing errors
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When a cell makes a protein it is said
that that gene is expressed.
Nearly all the body’s cells can make all
human proteins, but each type of cell
makes only the kinds of proteins it
needs.
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Roles of Proteins

Building material
 Growth, a matrix of protein underlies
almost all structures in the body including
bones,muscles ligaments, tendons,
connecting matrix between cell walls, scar
tissue, hair and nails.
 Maintenance, GI tract cells are replaced
every three days. The whole body has its
cells renewed every seven years.
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Proteins as Enzymes
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Enzymes are usually composed of a protein, a
vitamin and a trace mineral.
They act as catalysts, allowing reactions to
occur more quickly and efficiently.
They can cause two substances to come
together making a new structure or can split a
compound apart.
An enzyme is not affected by the chemical
reactions it allows to take place.
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Roles of Proteins

Enzymes
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Hormones
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Roles of Proteins

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Regulation of fluid balance
 Dependent edema may be caused when there is too
much fluid between cells and not enough
hydrophilic protein within the cells.
Acid-base regulation, proteins act as buffers
accepting and releasing hydrogen ions thus preventing
acidosis or alkalosis.
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Transport Proteins
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Proteins in Immunity


Antibodies are giant proteins that bind up specific
invaders like viruses or antigens
Antigens are substances that cause the body to
produce antibodies. They may include bacteria,
allergens, toxins or anything that causes an
inflammatory response.
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Roles of Proteins


Source of energy. The brain and nervous system must
have glucose. Once the amino group is removed from
the protein, the remaining carbon molecules can be
used to create energy - 4 Kcal per gram or stored as
fat.
Other roles include being converted to other proteins
or making neurotransmitters norepinephrine and
epinephrine, melanin, fibrin and as precursor to the
vitamin niacin.
Aulani "Biokimia" Presentation 5
Roles of Proteins
 Other roles include being converted
to
other proteins or making:
 neurotransmitters norepinephrine and
epinephrine
 melanin
 fibrin
 precursor to the vitamin niacin.
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Protein Metabolism

Protein turnover
 Amino acid pool includes free amino acids from
endogenous or exogenous sources
Nitrogen balance
 Positive nitrogen balance during growth or when
building new tissue
 Negative nitrogen balance if burned, fever, injury,
infection or starvation.
•
Aulani "Biokimia" Presentation 5
Protein Metabolism

–
Using amino acids to make proteins.
Cells can dismantle one amino acid and combine the
amino group of that amino acid with carbon fragments
from glucose metabolism to make another essential or
nonessential amino acid needed.
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Protein Metabolism


Deamination is the removal of the nitrogen containing
amino group, converting it to ammonia, which is sent
to the liver and converted into urea. The remaining
carbon fragment may be burned or stored as fat.
Amino acids can be used to make fat by removing the
amino group and converting the remaining carbon
fragments to fat.
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Amino Acid Pool
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
Proteins are degraded and resynthesized continuously
Several times more protein is turned over daily within
the body (endogenous) than is consumed (exogenous)
AA consumed in excess or unable to be used are not
stored. They are:
 degraded into urea, uric acid, and creatinine
 lost in feces or sweat
 converted into hair and nails
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Protein Quality
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High-quality proteins
Digestibility
 Animal vs. plant
Amino acid composition
 Limiting amino acid
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Protein Quality
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
Is reflected in the amino acid score:
content of
individual essential AA in food
 content of same AA in reference pattern
Based on reference pattern for age
Four AA are likely to be limiting
 Lysine, sulfur containing (methionine plus cystine),
threonine and tryptophan
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Protein Quality
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Reference protein
Complementary proteins
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Reference Proteins
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Nitrogen balance studies within age groups
Used data for highy digestible, high quality proteins egg, meat, milk or fish
Amino acid scoring patterns were factored in
A margin of safety of 2 standard deviations to meet
needs of 97.5% of the population
For adults over age 19 the reference protein intake is
0.75 g/kg/day (RDA is 0.8)
Range was 0.54 - egg to 0.99 - vegetable based diet
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Protein Quality
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Protein Digestibility Corrected AA Score
compares the amino acid content of a protein
with the human amino acid requirements and
corrects for digestibility.
Considers factors that limit digestion:
 cell walls, enzyme inhibitors, tannins
Reveals the most limiting AA
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Protein Quality

PDCAAS
 Protein digestibility-corrected
amino acid score
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Food Labels


Quantity of protein
Daily Value
 50 g protein
 10% of a 2000 kcal diet
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Protein-Energy Malnutrition
(PEM)


Acute PEM when one is recently deprived of food.
Children are thin for their height.
Chronic PEM from long term food deprivation.
Children are short for their age.
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Protein-Energy Malnutrition
Marasmus - inadequate energy and protein over a long
period of time. Often seen at 6-18 months of age.
Look like little old people.
• Kwashiorkor - “the evil spirit that infects the first
child when the second child is born.”
Sudden
deprivation at 18 mon to 2 yrs.
 Marasmus-kwashiorkor mix: edema of marasmus with
wasting of kwashiorkor

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Protein-Energy Malnutrition


Infections: Antibodies needed to fight
infections are degraded to provide amino
acids for survival.
 Dysentery with concomitant diarrhea robs
the body of needed nutrients.
Rehabilitation: electrolytes especially
potassium and sodium are given slowly over
the first 2 days, then foods may be started in
small quantities.
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Health Effects of High Protein

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
Heart disease: diet high in animal protein may
contribute to a higher incidence of heart disease in the
U.S. High homocysteine level possibly from suboptimal
B12, B6 and/or folic acid is associated with heart
disease.
Cancer of the colon, breast, kidneys, pancreas and
prostate is associated with high animal protein and fat
diet.
Adult bone loss. Calcium excretion rises as protein
intake increases.
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Health Effects of High Protein Diet



Weight control helpful for some not all
Any diet in which grains are severely limited
should supplement with manganese 5 mg/day and
selenium 200 ug
Kidney disease - a high protein diet increases
the load on the kidneys, which in Chinese
medicine means degenerative changes will take
place earlier. We are as old as our back is
flexible and our kidneys function adequately.
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Recommended Intakes

RDA
 0.8 g/kg/day for healthy adults
 8-11% - of energy intake per day
 50-65 grams of protein per day or between 200 to 250 Kcal.
 If junk food, sugar and fat is
restricted, it is
difficult not to get enough protein.
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Recommended Intakes
•

Calculate own: wt (lbs) divided by 2.2 x .8
Adequate intake - if total Calories are too low,
protein will be used to meet Calorie needs

Protein in abundance
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Supplements
•

Protein supplements- very active athletes may benefit
from an intake of 1 gm of protein per kg rather than .8
gm/kg of body wt. (75 gm instead of 63 gm/day - 1 1/2
oz more meat)
Amino acid supplements
 if not balanced may lead to deficiencies of some AA
thru competition with carrier enzymes
 Lysine up to 3 gm a day may suppress herpes
infection (divided doses with meals)
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Vegetarian Diets

Healthy food choices
 Macrobiotic diets use too much salt and can
be overly restrictive.
However, they
contribute one excellent idea - to eat as
much as possible a variety of minimally
processed, organic, locally grown foods in
the season in which they are grown.
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