Chapter 4 Amino Acids

General Properties

• There are 20 standard amino acids. With exception of proline, all have
primary amino group and a carboxylic acid bonded to same carbon.
• Carboxyl group and amino group all have pKa values around 2.2 and 9.4
respectively
• Note that at physiological pH of 7, amino acids are charged as shown to
left. They can act as either acid or base.
• Also referred to as zwitterions
• This imparts special properties e.g. some have melting points of 300 C,
just like salts. Also very soluble in water and not organics.
Peptide Bonds
• Peptide bond formed from
elimination of water from 2
amino acids resulting in CO-
NH linkage.
• Can have di-, tri-, oligo- and
poly-peptides.
• Proteins composed of linear
polymer of amino acids.
• Large number of proteins
can exist. A small protein of
100 amino acids can have
20100 or 1.27x10130 possible
unique polypeptide chains.
Classification of amino acids
• Side chains of amino acids are responsible
for many of the unique properties of
proteins
• 3 major classes of sides chains
– hydrophobic
– charged
– polar
– can also count 4 classes if Gly considered in own class
• The 20 different amino acids are
commonly abbreviated with 1 or 3-letter
codes
Hydrophobic Amino Acids 1
Hydrophobic Amino Acids 2
Charged Amino Acids
Polar Amino Acids 1
Polar Amino Acids 2
Hydrophobic Amino Acids 1
Disulfide-linked Cys residues
• Two adjacent cysteine
residues can be
oxidized to form a
disulfide bond
• disulfide bonds are
usually found in
extracellular and not
intracellular proteins
– inside of cell is a
reducing
environment
• disulfide bonds can
stabilize protein
structure by providing
crosslink
 Significance of Side Chains
• Select number can participate in
“chemistry” of enzyme active sites. Those
with charged or polar groups provide
interesting catalytic groups.
• Function of nonpolar amino acids? A
protein folding issue. Can think of protein
as hydrophobic core made of different
Lego pieces. These pieces have to fit
together in compact fashion.
Acid-Base Properties of Amino Acids
• Titration curve for glycine
– Note that one starts with
all groups in acid form.
– Note how many
equivalents are added
– Note that at 0.5 and 1.5
equivalents, pH is equal to
pK of group being
titrated.
– Note pH which gives zero
charge is the isoelectric
point. Calculated as (pK1+
pK2)/2
– Note where the buffering
capacity is best
More Titration Curves
 Environment and pKas
• pKa are influenced by environment.
• To predict influence of environment, examine the charge of
the acid and the conjugate base. E.g. His side chain has a
cationic acid and neutral base whereas Asp has neutral acid
and anionic base.
• Do groups near the acid stabilize the acid or the base. E.g.
if negative charge near a His residue, it would stabilize the
positive charge of the acid, thereby making it a weaker acid
and increasing the pK. Local environment can be:
– Positively charged
– Negatively charged
– Hydrophobic
• Based on above, explain why pK of a-carboxyl and a-amino
group is lower than expected.
 Misc. about proteins and nomenclature
• Proteins have complex titration curves.
Many ionizable side chains whose pKs can
be shifted by local environment
• Isoelectric point of proteins can be
calculated based on amino acid sequence
or experimentally determined with
isoelectric focusing.
• Assignment: determine the pI of a
hexapeptide: Gly-Glu-Asp-His-Lys-Ala
 Optical Activity

• All amino acids except for gly rotate

polarized light
• Optically active molecules have
asymmetry such that mirror images
are not superimposable
• Terms: asymmetic centers, chiral
centers, enantiomers.
• Enantiomers physically and
chemically indistinguishable by most
techniques
 Systems for describing enantiomers

• Direction which a cpd rotates

polarized light. Dextrorotator
(rt) also (+) or levorotatory (lf)
also (-). Determined by
polarimeter.
• Fischer convention is related to •The RS system depends on atomic
glyceraldehyde and uses D or L. numbers. Lowers is placed behind
• All amino acids from proteins chiral center. Number then from
have L stereochemical highest to lowest. R is clockwise
configuration. and S is counter clockwise.
 Diastereomers

L-threonine is 2S-3R, name the other diasteromers

 Spectroscopy of amino acids

Only the aromatic amino acids absorb light in the UV region

 Determination of extinction coefficient for a protein
• Variety of methods available, this one is useful when the sequence
is known
• Edelhoch et al. [Biochemistry (1967) 6, 1948-1954]
• From known sequence, determine Trp and Tyr content
• With purified protein of interest, determine the absorbance at 280
nm (or maximum near this wavelength) in buffered solution
• Take sample of known absorbance, dissolve in 6 M guanidine
hydrochloride in 0.02 M phosphate, pH 6.5. Record absorbance at
288 and 280.
• Use 2 simultaneous equations to determine the number of Trp and
Tyr:
– Abs. (288nm)= NTrp4815 + MTyr385
– Abs. (280) = NTrp5690 + MTyr1280
• Calculate concentration of Trp and Tyr. Divide by number of Trp
and Tyr per enzyme to obtain concentration of enzyme in solution.
 Prochiral

• Prochiral: replacement of one of the substituents

turns achiral center into a chiral one. Enzymes, like
you, can recognize and distinguish between pro R
and pro S atoms.
 More prochiral
H
H • Planar molecules can also have
C
prochiral nature.
C
O CH3
Acetaldehyde has a re face
H3C O and si face. Reduction of one
re face
si face
side versus the other yields a
different stereoisomer (if
using deuterium or tritium).
• Enzymes (dehydrogenases)
have different preferences
H H for re or si face of substrates
C
OH HO
C • For example, if the aldehyde
H3C D CH3 is reduced with deuterium
D
based on the re or si side, the
two different enantiomer
products are formed.
 Chirality and life
• Ordinary synthesis of chiral molecules
produces racemic mixtures. Ordinary
methods do not show stereochemical
preference.
• A fact of life: biosynthesis of substances
having asymmetric centers almost produce
pure stereoisomers.
• Using this criteria, examination of amino
acids in meterorites always show racemic
mixture. Thus not based on life.
• Not known why life has shown this
preference.