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Amino Acids
“When you understand the amino acids,
you understand everything”
©CMBI 2001
Amino Acids
Proteins are macromolecules made up from 20 different amino acids.
The heart of the amino acid is the so-called C. To which are bound: an
amino group, a carboxyl group, a hydrogen, and the side chain.
O
H2N
C
CH

OH
R
The C, C, N and O atoms are called backbone atoms.
R denotes any one of the 20 possible side chains.
©CMBI 2001
Amino Acids
Backbones run from the amino to the carboxy end, or in other
words, from N-terminus to C-terminus.
©CMBI 2001
Zwitterion
At pH 7, the amino group and the carboxyl groups are both ionised.
In this state the amino group is
protonated and thus positive, and
the carboxyl group is de-protonated,
and thus negative.
O
H3N
An amino acid in this polarised
state is called a zwitterion.
+
C
CH

O
R
©CMBI 2001
-
Di-peptide
Amino acids bind, to form a
protein. Upon binding, two
protons from the NH3 and
one oxygen from the
carboxyl join to form a water.
So the peptide bond has at
the one side a C=O and at
the other side an N-H.
Only the ends of the chain
are NH3 or carboxylic.
©CMBI 2001
The Peptide Bond
H2N
f
CH
R1
phi ()
psi ()
omega ()

O
R2
C
CH

OH
N
C
H
O
= torsion angle around N-CA bond
= torsion angle around CA-C bond
= connection between two amino acids
= peptide bond (always close to 180 degrees,
sometimes close to zero)
©CMBI 2001
20 Amino Acids
The side chains, R, determine the differences in the structural and
chemical properties of the 20 ‘natural’ amino acids.
The 20 amino acids can be classified as follows:
Aliphatic/hydrophobic
Polar
Alcoholic
Sulfur-containing
Aromatic
Charged
Special
Ala, Leu, Ile, Val
Asn, Gln
Ser, Thr, (Tyr)
Met, Cys
Phe, Tyr, Trp, (His)
Arg, Lys, Asp, Glu, (His)
Gly (no R), Pro (cyclic, imino-acid)
Several amino acids belong in more than one category.
©CMBI 2001
Amino Acid Sequence
The amino acid sequence (also called primary structure) of a
protein is the order of the amino acids in the protein chain.
The sequence is always read from the N-terminus to the Cterminus of the protein.
For example:
+H3N-Lys-Val-Phe-Ala-Met-Cys-Leu-Leu-Arg-Val-COO-
Or (in one-lettercode):
KVFAMCLLRV
©CMBI 2001
Direction of Protein Chain
Lys-Val
Val-Lys
©CMBI 2001
The 20 Amino Acids
A
C
D
E
F
G
H
I
K
L
M
N
P
Q
R
S
T
V
W
Y
Ala
Cys
Asp
Glu
Phe
Gly
His
Ile
Lys
Leu
Met
Asn
Pro
Gln
Arg
Ser
Thr
Val
Trp
Tyr
Alanine
Cysteine
Aspartic acid (Aspartate)
Glutamic acid (Glutamate)
Phenylalanine
Glycine
Histidine
Isoleucine
Lysine
Leucine
Methionine
Asparagine
Proline
Glutamine
Arginine
Serine
Threonine
Valine
Tryptophan
Tyrosine
©CMBI 2001
The 20 Amino Acids
©CMBI 2001
Amino Acid Characteristics
There are many of ways to characterize the properties of amino
acids. The ones most useful and most commonly used are:
Hydrophobicity
Size
Charge
Secondary structure preference
Alcoholicity
Aromaticity
And on top of that there are some special characteristics like bridge
forming by cysteines, rigidity of prolines, titrating at physiological
pH of histidine, flexibility of glycines, etc.
©CMBI 2001
Hydrophobicity
Hydrophobicity is the most important characteristic of amino acids.
It is the hydrophobic effect that drives proteins towards folding.
Actually, it is all done by water. Water does not like hydrophobic
surfaces. When a protein folds, exposed hydrophobic side chains
get buried, and release water of its sad duty to sit against the
hydrophobic surfaces of these side chains.
Water is very happy in bulk water because there it has on average
3.6 H-bonds and about six degrees of freedom.
So, whenever we discuss protein structure, folding, and stability, it
is all the entropy of water, and that is called the hydrophobic effect.
©CMBI 2001
Secondary Structure Preference
Amino acids form chains, the sequence or primary structure.
These chains fold in -helices, b-strands, b-turns, and loops (or for
short, helix, strand, turn and loop), the secondary structure.
These secondary structure elements fold further to make whole
proteins, but more about that later.
There are relations between the physico-chemical characteristics of
the amino acids and their secondary structure preference. I.e., the
b- branched residues (Ile, Thr, Val) like to sit in b-strands.
We will now discuss the 20 ‘natural’ amino acids, and we will later
return to the problem of secondary structure preferences.
©CMBI 2001
A Ala Alanine
Alanine is a small, hydrophobic
residue. Its side chain, R, is
just a methyl group. Alanine
likes to sit in an alpha helix,it
doesn’t like beta strand very
much, but it hates beta-turns.
If you want to mutate a
residue, but you don’t have a
good plan about what to
replace it with, take alanine.
©CMBI 2001
C Cys Cysteine
Cysteine is a small till
intermediately large
hydrophobic residue. It doesn’t
like the alpha helix, but doesn’t
mind strand and turn. It can
form Cys-Cys bridges with
other cysteines. It can bind
metals (especially Zn and Cu).
The S-H group is very reactive.
©CMBI 2001
D Asp Aspartic acid
Aspartic acid, or aspartate, is
an intermediately large,
hydrophilic, negatively charged
residue. Its side chain normally
titrates at pH 4.5. It likes to sit
near the N-terminus of a helix,
and in turns. It hates strands.It
often occurs in active sites. It
can bind ions (mainly Ca).
©CMBI 2001
E Glu Glutamic acid
Glutamic acid, or glutamate, is
a large, hydrophilic, negatively
charged residue. Its side chain
titrates at pH 4.6. It loves the
helix, doesn’t mind being in a
strand, but is not so good for a
turn.
©CMBI 2001
F Phe Phenylalanine
Phenylalanine is a large,
hydrophobic, aromatic residue.
It is good for a strand, it
doesn’t mind sitting in a helix,
but it hates the turn.
©CMBI 2001
G Gly Glycine
Glycine is the smallest residue.
It doesn’t have a side chain, so
its hydrophobicity is a bit
undetermined, but you can call
it hydrophobic or assign it an
intermediate hydrophobicity.
The fact that it doesn’t have a
side chain means that its
backbone is very flexible so
that it can make backbone
turns that other residues cannot
make. It is very bad for helix,
bad for strand, but it is the star of the turns.
©CMBI 2001
H His Histidine
Histidine is very special. It is
hydrophilic due to the two
nitrogens in its side chain. Both
nitrogens can titrate (the first
one at pH 6.2). It is a little bit
aromatic. It is often seen in
active sites.It is neutral at
physiological pH, but it can
easily become positive, and
occasionally even negative. It
can bind metal ions (mainly Zn,
Ni, Cu). It is not particularly
picky about its secondary structure.
©CMBI 2001
I Ile Isoleucine
Isoleucine is an intermediately
large, hydrophobic residue. It
is beta branched which means
that it likes to sit in a strand. It
doesn’t mind sitting in a helix
either, but it cries its eyes out
in a turn.
©CMBI 2001
K Lys Lysine
Lysine is a large, hydrophilic,
positively charged residue. It
is not a good strand residue,
but it doesn’t mind sitting in a
helix or in a turn. Its side chain
is very long and flexible.
©CMBI 2001
L Leu Leucine
Leucine is an intermediately
large, hydrophobic residue
that really loves to sit in a helix.
It is also good for a strand, but
it hates turns.
©CMBI 2001
M Met Methionine
Methionine is a large, sulphur
containing, hydrophobic
residue. It loves helices,
doesn’t mind sitting in a strand,
but it hates turns. Methionine
can bind metals with its sulphur,
but this sulphur is much less
reactive than the sulphur in
cysteine. It is often the first
residue of a molecule. The
N-terminus is mostly positive
and thus mostly at the surface.
Therefore, methionine, despite
being hydrophobic, is often at the surface.
We call this a forced marriage.
©CMBI 2001
N Asn Asparagine
Asparagine is an intermediately
large, polar residue. It hates the
helix, is mildly un-amused in a
strand, but it loves the turn. It
can bind metal ions (Ca), but
doesn’t do that as well as its
isosteric partner aspartic acid.
©CMBI 2001
P Pro Proline
In proline, the side chain is
connected to the backbone at
two places: the C and the N.
Therefore, it is not an amino
acid, but an imino acid. Unless
it is the N-terminal residue,
proline does not have a
backbone proton, and thus is
not good for helices and
strands. Due to the extra
covalent bond, proline is
already pre-bend, and thus
good for turns. Even though it is very hydrophobic, it often sits at
the surface
©CMBI 2001
Q Gln Glutamine
Glutamine is a large, polar
residue. It is not very picky
about its secondary structure.
It is isosteric with glutamic acid.
©CMBI 2001
R Arg Arginine
Arginine is a big, hydrophilic,
positively charged residue. Its
side chain contains a so-called
guadinium group that is rigid.
It is not picky about its
secondary structure.
©CMBI 2001
S Ser Serine
Serine is a small, alcoholic
residue of intermediate
hydrophobicity. It is not too
happy in helices and strands,
but it loves to sit in turns. It
often forms the active site of
an enzyme together with
histidine and aspartic acid. It
is occasionally involved in
metal (Ca) binding.
©CMBI 2001
T Thr Threonine
Threonine is a small, alcoholic
residue of intermediate
hydrophobicity. It is betabranched and thus good for
beta strands. It doesn’t care
about helices or turns. It is
occasionally involved in metal
(Ca) binding.
©CMBI 2001
V Val Valine
Valine is a small hydrophobic
residue. It is beta-branched
and thus good for beta strands.
It is isosteric with threonine.
Valine doesn’t care about
helices, but it hates turns.
©CMBI 2001
W Trp Tryptophan
Tryptophan is the biggest
residue. It is aromatic. Despite
that the nitrogen in the five-ring
is donor for hydrogen bonds, it
is very hydrophobic. It doesn’t
care about helices or turns, but
it loves strands.
©CMBI 2001
Y Tyr Tyrosine
Tyrosine is a large, aromatic,
alcoholic residue of
intermediate hydrophobicity. It
is not so happy in a helix,
indifferent about turns, and it
loves a strand.
©CMBI 2001
Secondary Structure Preferences
Alanine
Arginine
Aspartic Acid
Asparagine
Cysteine
Glutamic Acid
Glutamine
Glycine
Histidine
Isoleucine
Leucine
Lysine
Methionine
Phenylalanine
Proline
Serine
Threonine
Tryptophan
Tyrosine
Valine
helix
1.42
0.98
1.01
0.67
0.70
1.39
1.11
0.57
1.00
1.08
1.41
1.14
1.45
1.13
0.57
0.77
0.83
1.08
0.69
1.06
strand
0.83
0.93
0.54
0.89
1.19
1.17
1.10
0.75
0.87
1.60
1.30
0.74
1.05
1.38
0.55
0.75
1.19
1.37
1.47
1.70
turn
0.66
0.95
1.46
1.56
1.19
0.74
0.98
1.56
0.95
0.47
0.59
1.01
0.60
0.60
1.52
1.43
0.96
0.96
1.14
0.50
©CMBI 2001
Secondary Structure Preferences
Alanine
Glutamic Acid
Glutamine
Leucine
Lysine
Methionine
Phenylalanine
helix
1.42
1.39
1.11
1.41
1.14
1.45
1.13
strand
0.83
1.17
1.10
1.30
0.74
1.05
1.38
turn
0.66
0.74
0.98
0.59
1.01
0.60
0.60
Subset of helix-lovers. If we forget alanine (I don’t understand that
things affair with the helix at all), they share the presence of a
(hydrophobic) C-b, C-g and C-d (S-d in Met). These hydrophobic
atoms pack on top of each other in the helix. That creates a
hydrophobic effect.
©CMBI 2001
Secondary Structure Preferences
Isoleucine
Leucine
Phenylalanine
Threonine
Tryptophan
Tyrosine
Valine
helix
1.08
1.41
1.13
0.83
1.08
0.69
1.06
strand
1.60
1.30
1.38
1.19
1.37
1.47
1.70
turn
0.47
0.59
0.60
0.96
0.96
1.14
0.50
Subset of strand-lovers. These residues either have in common
their b-branched nature (Ile, Thr, Val) or their hydrophobicity (rest).
©CMBI 2001
Secondary Structure Preferences
Aspartic Acid
Asparagine
Glycine
Proline
Serine
helix
1.01
0.67
0.57
0.57
0.77
strand
0.54
0.89
0.75
0.55
0.75
turn
1.46
1.56
1.56
1.52
1.43
Subset of turn-lovers. Glycine is special because it is so flexible, so
it can easily make the sharp turns and bends needed in a b-turn.
Proline is special because it is so rigid; you could say that it is prebend for the b-turn.
Aspartic acid, asparagine, and serine have in common that they
have short side chains that can form hydrogen bonds with the own
backbone. These hydrogen bonds compensate the energy loss
caused by bending the chain into a b-turn.
©CMBI 2001
Sequence Alignment
Now that we know everything about the amino acids, we can look
at the real goal of this whole excercise: sequence alignments.
Which of the following alignments is better, left or right:
CWPSAAFPWC
CWPT---PWC
CWPSAAFPWC
CWP---TPWC
CWPYAAWPWC
CWP---FPWC
CWPYAAWPWC
CWPF---PWC
©CMBI 2001
Sequence Alignment
Don’t forget that we still want to gather information about an
unknown protein for which we determined the sequence.
To gather that information, we will need databases and sequence
alignments.
To do these sequence alignments, we need to know everything
about the amino acids.
And that is what we are working on right now.
©CMBI 2001