Chapter 26 Amino Acids, Peptides, Proteins

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Chapter 26Amino Acids, Peptides, Proteins
Structure an amino acid possesses both a
carboxyl and an amino group. The most common in
the biological amino acids are the a-amino acids.
The amino acid exists as a dipolar salt,a
zwitterion, possessing both a positive charge and
a negative charge.
zwitterion
Amino acids have high mps, are soluble in water,
and insoluble in nonpolar organic solvents.
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Chirality
With the exception of glycine, all
protein-derived amino acids have at least one
stereocenter and are therefore chiral
D-alanine L-alanine
The vast majority of amino acids in the
biological world are the L series
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Protein-derived Amino Acids
Nonpolar side chains
Alanine Glycine Isoleucine
Leucine
Methionine Phenylalanine
Proline
Tryptophan
Valine
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Polar Side Chains
Asparganine Serine Glutamin
e Threonine
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Acidic Side Chains
Aspartic acid Glutamic acid Arginine
Histidine Tyrosine
Lysine
Cysteine
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Some Characteristics of Protein-derived Amino
Acids

• All 20 are a-amino acids, meaning that the amino
  group is located on the carbon alpha to the
  carboxyl group.
• 2. For 19 of 20, the a-amino group is primary.
  Proline is different.
• 3. Except for glycine, the a-carbon is a
  stereocenter. All 19 are L amino acids
  (S)-amino acids.
• 4. Isoleucine and threonine contain a second
  stereocenter. Four stereoisomers are possible,
  but only one is found in proteins.

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Acid-Base Properties of Amino Acids
Acidity of a-carboxyl group
The average pKa of the carboxyl group is 2.19,
considerably Stronger than that of acetic acid
(pKa 4.76).
pKa 2.2
Ammonium group is electron-withdrawing by
induction and stabilizes carboxylate anion.
pKa 4.76
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Acidity of a-ammonium groups
pKa 9.5
pKa 10.60
a-Ammonium group is slightly stronger acid than
that of primary ammonium group. Conversely,
a-amino group is slightly stronger base than
primary amino group.
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Isoelectric Point (pI) The pH at which an
amino acid, polypeptide or protein has no
net charge (zwitterion).
pI for glycine pI ½(pKa a-COOH pKa
a-NH3) ½ (2.35 9.78) 6.06
At pH 6.06, the predominant form of glycine is
the dipolar ion or zwitterion, H3NCH2CO2-
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Amino Acid Analysis Capillary Electrophoresis/Fluo
rescent Detection
NDA aa
NDA-aa Fluorescence
labs 420 nm lemisson 490 nm
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Polypeptides and Proteins
In 1902, Emil Fischer proposed that proteins are
long-chains of amino acids joined together by
amide bonds.
Peptide bond
By convention, N-terminal amino acid on left and
C-terminal amino acid on right
Serylalanine (Ser-Ala,S-A)
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Problem Draw a structural formula of
Cys-Arg-Met-Asn. Label theN-terminal amino acid
and the C-terminal amino acid. What is the net
charge on the peptide at pH 6.0?
pKa 8.00
C-terminal
N-terminal
pKa 12.48
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Primary Structure of Polypeptides and Proteins
Primary structure (1o) refers to sequence of
amino acids. How can this be determined?

• Amino acid analysis. Hydroysis using 6 M HCl in
  sealed
• glass tubes at 110oC for 24 hours. Mixture is
  separated by
• ion exchange chromatography. Ninhydrin
  derivatizes the
• amino acids and absorbance is measured.
• Sequence Analysis.
• a. Cyanogen bromide cleavage
• b. Enzyme-catalyzed Hydroysis
• c. Edman degradation
• d. Mass spectrometry

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• Cyanogen bromide cleavage specifically cleaves
• peptide bonds formed from carboxyl group of
  methionine.

Bond is cleaved
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2. Enzyme catalyzed Hydrolysis of Peptide Bonds
Trypsin catalyzes the hydrolytic cleavage of
peptide bonds Formed by the carboxyl groups of
arginine and lysine.
arginine
lysine
Chymotripsin - catalyzes the hydrolytic cleavage
of peptide bondsformed by the carboxyl groups of
phenylalanine, tyrosine, and tryptophan
phenylalanine tryptophan tyrosine
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Problem Which of these dipeptides is hydrolyzed
by trypsin? By chymotripsin?
a. Arg-Glu-Ser b. Phe-Gly-Lys

trypsin Answer Arg-Glu-Ser H2O
Arg Glu-Ser
chymotrypsin

Phe-Gly-Lys H2O Phe
Gly-Lys
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3. Edman Degradation most widely used
N-terminal amino acid
Phenyl isothiocyanate
Phenylthiohydantoin separated and identified.
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Adsorbed onto solid surface
Nucleophilic acyl addition to isothiocyanate
Nucleophilic acyl substitution of amide
Washed off and identified by liquid chromatography
rearrange
Phenylthiohydantoin