1. Proteins are polymers of amino acids.
There are 20 unique amino acids that make up proteins in general. Each of the 20 different amino acids has a different R group.
Otherwise, the amino acid structure is the same!
Generic structure of an amino acid
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2. KEY CONCEPTS: Section 4-1
The 20 amino acids differ in the chemical characteristics of their R groups.
Amino acids are linked by peptide bonds to form a polypeptide.
A protein’s structure may be described at four levels, from primary to quaternary.
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3. Amino
Acids:
Get to
Know
Them!!

4. Amino
Acids:
I’m not
Kidding.
Make
them
your
friends!!

5. © 2014 John Wiley & Sons, Inc. All rights reserved.
The 20 amino acids differ in the chemical characteristics of their R groups.
There are three categories for the R groups.
Hydrophobic amino acids have nonpolar R groups.
Hydrophilic amino acids have polar R groups.
Polar R groups
Nonpolar R groups
Uncharged
Charged
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6. Glycine has a nonpolar side chain.
Gly is the simplest amino acid.
Side chain = H
Each amino acid has:
Full name (e.g., Glycine)
3 letter code (e.g., Gly)
1 letter code (e.g., G)
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7. Ala and Phe have hydrophobic R groups.
Ala has a methyl group for its side chain.
Phe, as its name suggests, includes a phenyl ring on an Ala residue.
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8. Val, Leu and Ile have hydrophobic R groups.
Leu has an extra methylene group (-CH2-) than Val.
Ile has the same functional moieties in its R group as Leu, just arranged differently – hence the prefix iso.
V, L, and I are also referred to as Branched Chain Amino Acids (BCAAs)
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9. Met has a hydrophobic R group.
Met is one of only two amino acids with a sulfur in its R-group.
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10. Trp has a hydrophobic R group.
Trp is the only amino acid with a fused ring system.
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11. Practical Application
Most proteins have at least one Trp.
Trp absorbs UV light at 280 nm.
[Protein] can be determined based on detection of Trp in proteins!
Recall
Beer’s Law:
Aλ = ελbc
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12. © 2014 John Wiley & Sons, Inc. All rights reserved.
Pro has a unique R group.
Pro is the only amino acid whose side chain loops back onto its own backbone!
Pro induces kinks in a polypeptide sequence.
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13. Hydrophilic Amino Acids
There are 2 groups of hydrophilic amino acids.
Polar, uncharged
Polar, charged
Polar amino acids are often found in the active sites of enzymes because they can facilitate chemical catalysis.
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14. Cys residues can form disulfide bonds.
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15. Disulfide bonds facilitate crosslinking.
Disulfide bonds can form intra-strand crosslinks (as shown).
When a protein contains more than one polypeptide chain, disulfide bonds can also form inter-chain crosslinks.
3D Structure of the Protein Lysozyme
Only the backbone is shown in blue. Disulfide bonds are shown in yellow ball-and-stick representation.
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16. Ser and Thr have hydroxyl groups in their side chains.
-OH groups are prominent nucleophiles in biochemical reactions.
Other chemical groups can covalently bond to proteins via -OH groups.
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17. Tyr has a polar, uncharged R group.
Tyr is a derivative of Phe.
Phe and Tyr are precursors of amino acid derivatives that are neurotransmitters.
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18. Asn and Gln have polar, uncharged R groups.
Gln has 2 methylene groups, Asn has only 1.
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19. His has a polar, uncharged R group.
Histidine has an imidazole (5-membered ring system with N’s).
NOTE: Histidine can be charged below its pKa of ~6.
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20. Lys and Arg have positively charged, polar R groups.
Both Lys and Arg have long side chains with a positively charged amine group.
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21. Asp and Glu have negatively charged, polar R groups.
Asp and Glu are analogous to Asn and Gln, except Asp and Glu have carboxylate groups!
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22. Amino acids are linked via a condensation reaction.
Here amino acids within the peptide are called “amino acid residues” because only the residual atoms remain.
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23. Amino acids are linked by peptide bonds to form a polypeptide
Example below shows a short peptide.
Polypeptides have many amino acid residues.
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24. Solution: Look up the pK values for the ionization states.
Recall the meaning of pK!
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25. pK reveals the cutoff pH for protonation of a species.
When the pH < 3.5, the structure of a generic amino acid is
pK = 3.5
Both ends are protonated at very low pH.
pK = 9.0
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26. pK reveals the cutoff pH for protonation of a species.
When the pH > 3.5 and <9.0, the structure of a generic amino acid is
Carboxyl group is deprotonated at neutral pH.
Amino group is still protonated!
pK = 3.5
Zwitterion
Form of a Generic Amino Acid
pK = 9.0
© 2014 John Wiley & Sons, Inc. All rights reserved.

27. pK reveals the cutoff pH for protonation of a species.
When the pH > 9.0, the structure of a generic amino acid is
pK = 3.5
Carboxyl group is still deprotonated.
Amino group is now deprotonated!
pK = 9.0
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28. © 2014 John Wiley & Sons, Inc. All rights reserved.
PROBLEM: How can one deduce the structure of an amino acid or peptide when the side chain also has an ionizable group?
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29. Solution: Look up the pK values for the ionization states.
Recall the meaning of pK!
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30. Solution: Analyze side chain pK values.
EXAMPLE: His, pK = 6.0
At pH < 6.0, His side chain is protonated
At pH > 6.0, His side chain is deprotonated
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31. There are four different levels of protein structure.
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33. KEY CONCEPTS: Section 4-2
The polypeptide backbone has limited conformational flexibility.
The α helix and β sheet are common secondary structures characterized by hydrogen bonding between backbone groups.
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34. Biochemists view macromolecules in a variety of ways!
“Ribbon” diagram (red) Ball-and-stick atoms are superimposed
Hydrogen bonds = dashed lines
Space-filling representation of all atoms. Shades of blue = different subunits
Shows that proteins are globular in 3D!
Shows shape of backbone chain in one region of a protein is alpha helical!
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37. Hydrogen Bonding in a β Sheet
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38. Some polypeptide chains align in regions with directionality.
Antiparallel Beta Sheets align in opposite directions.
Parallel Beta Sheets align in the same direction.
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39. © 2014 John Wiley & Sons, Inc. All rights reserved.
Secondary Structures
The α helix and β sheet are common secondary structures characterized by hydrogen bonding between backbone groups.
H-bonds form in an α helix between the carbonyl oxygen and the amino hydrogen.
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41. Proteins can have any combination of secondary structures.
α-helical protein
β-protein
α/β protein
Protein with very little 2° structure
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42. KEY CONCEPTS: Section 4-3
A folded polypeptide assumes a shape with a hydrophilic surface and a hydrophobic core.
Protein folding and protein stabilization depend on noncovalent forces.
Some proteins can adopt more than one stable conformation.
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43. © 2014 John Wiley & Sons, Inc. All rights reserved.
A folded polypeptide assumes a shape with a hydrophilic surface and a hydrophobic core.
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44. © 2014 John Wiley & Sons, Inc. All rights reserved.
Protein folding and protein stabilization depend on noncovalent forces.
Key example: the hydrophobic effect
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45. KEY CONCEPTS: Section 4-4
Proteins containing more than one polypeptide chain have quaternary structure.
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46. A single chain can form local 3D structure – domains.
Domains vs. Subunits
Two or more separate chains (subunits) can orient in 3D space to give quaternary structure!
A single chain can form local 3D structure – domains.
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47. KEY CONCEPTS: Section 4-5
Chromatography is a technique for separating molecules on the basis of size, charge, or specific binding behavior.
The sequence of amino acids in a polypeptide can be determined.
The 3D arrangement of atoms in a protein can be deduced by measuring the diffraction of X-rays or by analyzing NMR.
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48. © 2014 John Wiley & Sons, Inc. All rights reserved.
Chromatography is a technique for separating molecules on the basis of size, charge, or specific binding.
Common chromatographic methods in biochemistry
Gel filtration (or size exclusion) chromatography
Ion exchange chromatography
Affinity chromatography
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49. Gel Filtration Chromatography
Separation based on size
Small Proteins
Large Proteins
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50. Ion Exchange Chromatography
Separation based on charge
Resins have either diethylaminoethane or carboxymethyl functional groups.
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51. SDS PAGE Analysis of Proteins

53. Isoelectric Points of Several Common Proteins.

54. 2D Gel Analysis - combines SDS-PAGE with Isoelectric Focusing

55. Crystals of the Protein Streptavidin
The 3D arrangement of atoms in a protein can be deduced by measuring the diffraction of X-rays or by analyzing NMR.
With X-ray crystallography, the protein must first be crystallized.
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56. The NMR Structure of Glutaredoxin
The 3D arrangement of atoms in a protein can be deduced by measuring the diffraction of X-rays or by analyzing NMR.
With NMR spectroscopy, a family of structures is obtained.
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