1. Proteins: Function & Structure

2. Proteins Cellular Overview Core Topics Functions Key Properties
Amino Acids: properties, classifications, pI
Primary Structure, Secondary Structure, and Motifs
Tertiary Structure
Fibrous vs. Globular
Quaternary Structure

3. Amazing Proteins: Function
Catalysts (Enzymes)
Transport & Storage
The largest class of proteins, accelerate rates of reactions
DNA Polymerase
Catalase
CK2 Kinase
Ovalbumin
Casein
Ion channels
Hemoglobin
Serum albumin

4. Amazing Proteins: Function
Structural
Generate Movement
Collagen
Keratin
Silk Fibroin
Actin
Myosin

5. Amazing Proteins: Function
Regulation of Metabolism and Gene Expression
Protection
Insulin
Lac repressor
Thrombin and Fibrinogen
Ricin
Immunoglobulins
Venom Proteins

6. Amazing Proteins: Function
Signaling and response (inter and intracellular)
Apoptosis
Membrane proteins
Signal transduction

7. Amazing Proteins: Properties
Biopolymers of amino acids
Contains a wide range of functional groups
Can interact with other proteins or other biological macromolecules to form complex assemblies
Some are rigid while others display limited flexibility

8. a-Amino Acids: Protein Building Blocks
R-group or side-chain
a-amino group
Carboxyl group
a-carbon

9. Amino acids are zwitterionic
“Zwitter” = “hybrid” in German
Fully protonated forms will have specific pKa’s for the different ionizable protons
Amino acids are amphoteric (both acid and base)

10. Stereochemistry of amino acids

11. Stereochemistry of amino acids (AA)
AA’s synthesized in the lab are racemic mixtures. AA’s from nature are “L” isomers
These are all optically-active except for glycine (why?)

12. Synthesis of Proteins
+ H2O

13. Synthesis of Proteins

14. Synthesis of Proteins
N-Terminal End
C-Terminal End

15. Synthesis of Proteins ≠ =

16. Synthesis of Proteins ≠ =

17. Synthesis of Proteins ≠ ≠

18. COMMON AMINO ACIDS
20 common amino acids make up the multitude of proteins we know of

19. Amino Acids With Aliphatic Side Chains

20. Amino Acids With Aliphatic Side Chains

21. Amino Acids With Aliphatic Side Chains

22. Amino Acids With Aromatic Side Chains

23. Amino Acids with Aromatic Side Chains Can Be Analyzed by UV Spectroscopy

24. Amino Acids With Hydroxyl Side Chains

25. Amino Acid with a Sulfhydryl Side Chain

26. Disulfide Bond Formation

27. Amino Acids With Basic Side Chains

28. Amino Acids With Acidic Side Chains and Their Amide Derivatives

30. There are some important uncommon amino acids

31. pH and Amino Acids
Net charge: +1
Net charge: 0
Net charge: -1

32. Characteristics of Acidic and Basic Amino Acids
Acidic amino acids
Low pKa
Negatively charged at physiological pH
Side chains with –COOH
Predominantly in unprotonated form
Basic amino acids
High pKa
Function as bases at physiological pH
Side chains with N

34. Isoelectic point (pI)
the pH at which the compound is electrically neutral
Equal number of (+) and (-) charge
At pH < pI amino acid is (+)
At pH > pI amino acid is (-)
CRITICAL FOR: protein analysis, purification, isolation, crystallization

38. Protein Structure
We use different “levels” to fully describe the structure of a protein.

39. Primary Structure Amino acid sequence
Standard: Left to Right means N to C-terminal
Eg. Insulin (AAA40590)
The info needed for further folding is contained in the 1o structure.
MAPWMHLLTVLALLALWGPNSVQAYSSQHLCGSNLVEALYMTCGRSGFYRPHDRRELEDLQVEQAELGLEAGGLQPSALEMILQKRGIVDQCCNNICTFNQLQNYCNVP

40. Secondary Structure
The regular local structure based on the hydrogen bonding pattern of the polypeptide backbone
α helices
β strands (β sheets)
Turns and Loops
WHY will there be localized folding and twisting? Are all conformations possible?

41. Consequences of the Amide Plane
Two degrees of freedom per residue for the peptide chain
Angle about the C(alpha)-N bond is denoted phi
Angle about the C(alpha)-C bond is denoted psi
The entire path of the peptide backbone is known if all phi and psi angles are specified
Some values of phi and psi are more likely than others.

42. The angles phi and psi are shown here
See blackboard for explanation why the peptide bond is planar

43. Unfavorable orbital overlap precludes some combinations of phi and psi
phi = 0, psi = 180 is unfavorable
phi = 180, psi = 0 is unfavorable
phi = 0, psi = 0 is unfavorable

44. Steric Constraints on phi & psi
Sasisekharan
G. N. Ramachandran was the first to demonstrate the convenience of plotting phi,psi combinations from known protein structures
The sterically favorable combinations are the basis for preferred secondary structures

45. α Helix First proposed by Linus Pauling and Robert Corey in 1951.
3.6 residues per turn, 1.5 Angstroms rise per residue
Residues face outward

46. α Helix α-helix is stabilized by H-bonding between CO and NH groups
Except for amino acid residues at the end of the α-helix, all main chain CO and NH are H-bonded

47. α Helix representation

49. β strand Fully extended
β sheets are formed by linking 2 or more strands by H-bonding
Beta-sheet also proposed by Corey and Pauling in 1951.

51. PARALLEL
ANTIPARALLEL

52. The Beta Turn (aka beta bend, tight turn)
allows the peptide chain to reverse direction
carbonyl C of one residue is H-bonded to the amide proton of a residue three residues away
proline and glycine are prevalent in beta turns

53. Mixed β Sheets

54. Twisted β Sheets

56.  Loops

57. What Determines the Secondary Structure?
The amino acid sequence determines the secondary structure
The α helix can be regarded as the default conformation
– Amino acids that favor α helices:
Glu, Gln, Met, Ala, Leu
– Amino acids that disrupt α helices:
Val, Thr, Ile, Ser, Asx, Pro

58. What Determines the Secondary Structure?
Branching at the β-carbon, such as in valine, destabilizes the α helix because of steric interactions
Ser, Asp, and Asn tend to disrupt α helices because their side chains compete for H-bonding with the main chain amide NH and carbonyl
Proline tends to disrupt both α helices and β sheets
Glycine readily fits in all structures thus it does not favor α helices in particular

59. Can the Secondary Structure Be Predicted?
Predictions of secondary structure of proteins adopted by a sequence of six or fewer residues have proved to be 60 to 70% accurate
Many protein chemists have tried to predict structure based on sequence
Chou-Fasman: each amino acid is assigned a "propensity" for forming helices or sheets
Chou-Fasman is only modestly successful and doesn't predict how sheets and helices arrange
George Rose may be much closer to solving the problem. See Proteins 22, (1995)

61. Modeling protein folding with Linus (George Rose)

62. Tertiary Structure The overall 3-D fold of the polypeptide chain
The amino acid sequence determines the tertiary structure (Christian Anfinsen)
The polypeptide chain folds so that its hydrophobic side chains are buried and its polar charged chains are on the surface
Exception : membrane proteins
Reverse : hydrophobic out, hydrophilic in
A single polypeptide chain may have several folding domains
Stabilized by H-bonding, LDF, noncovalent interactions, dipole interactions, ionic interactions, disulfide bonds

63. Fibrous and Globular Proteins

64. Fibrous Proteins
Much or most of the polypeptide chain is organized approximately parallel to a single axis
Fibrous proteins are often mechanically strong
Fibrous proteins are usually insoluble
Usually play a structural role in nature

65. Examples of Fibrous Proteins
Alpha Keratin: hair, nails, claws, horns, beaks
Beta Keratin: silk fibers (alternating Gly-Ala-Ser)

66. Examples of Fibrous Proteins
Collagen: connective tissue- tendons, cartilage, bones, teeth
Nearly one residue out of three is Gly
Proline content is unusually high
Unusual amino acids found: (4-hydroxyproline, 3- hydroxyproline , 5- hydroxylysine)
Special uncommon triple helix!

67. Globular Proteins
Most polar residues face the outside of the protein and interact with solvent
Most hydrophobic residues face the interior of the protein and interact with each other
Packing of residues is close but empty spaces exist in the form of small cavities
Helices and sheets often pack in layers
Hydrophobic residues are sandwiched between the layers
Outside layers are covered with mostly polar residues that interact favorably with solvent

68. An amphiphilic helix in flavodoxin:
A nonpolar helix in citrate synthase:
A polar helix in calmodulin:

70. Quaternary Structures
Spatial arrangement of subunits and the nature of their interactions. Can be hetero and/or homosubunits
Simplest example: dimer (e.g. insulin)
ADVANTAGES of 4o Structures
Stability: reduction of surface to volume ratio
Genetic economy and efficiency
Bringing catalytic sites together
Cooperativity

72. Protein Folding CHAPERONES assist protein folding
The largest favorable contribution to folding is the entropy term for the interaction of nonpolar residues with the solvent
CHAPERONES assist protein folding
to protect nascent proteins from the concentrated protein matrix in the cell and perhaps to accelerate slow steps