The Three Dimensional Structure of Proteins

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					           Chapter 6
The Three-Dimensional Structure
          of Proteins
            1, 9,12,
  Proteins: Higher Orders of Structure
• The structural variety of human proteins
  reflects the sophistication and diversity of
  their biologic role

• The maturity of a newly synthesized
  polypeptide into a biologically active protein
  requires that it be folded into a specific 3-D
  arrangement or conformation
  Conformation vs. Configuration
• Configuration deals with the arrangement of
  specific bonds about individual atoms

• Conformation refers to the spatial relationship
  of every atom of a molecule

• Proteins were initially characterized by their
  gross characteristics
         Initial Characterizations
•   Soluble proteins
•   Globular proteins
•   Fibrous proteins
•   Lipoproteins
•   Glycoproteins
•   Metalloproteins
Currently classification and Folding
• Now, proteins are classified based on
  similarity or Homology, of residue sequence
  and structure

• Typical proteins could have >1050 possible
• But since they fold as they form, it is easier to
  achieve biologically active conformation
      Orders of protein structure
• Primary Structure, 1o

• Secondary Structure, 2o

• Tertiary Structure, 3o

• Quaternary Structure, 4o
         Secondary Structure
• The number of possible secondary structures
  is restricted
• The is only free rotation about 2 of the 3
  bonds in the backbone
• Phi angle
• Psi angle
                Alpha Helix
• Figure 6.3a, page 164
• Both Phi and Psi angles are defined as well as
  distance it rises per turn
• Complete turn averages 3.6 residues
• R groups face outwards
• Mostly only righthanded a-Helix seen in
  nature due to mostly L-amino acids used!
          Stabilizing the helix
• H-bonds
• van der Waals in core
• Proline is only found in 1st turn. Why?
• When present elsewhere, disrupts helix and
  forms a bend
• Most hydrophobic R groups are on one side
  and hydrophilic on the other. Effect?
             The Beta Sheet
• As opposed to the helix, the amino acids are
  extended to form a zigzag or pleated pattern
• The R groups alternate up/down for a joining
• Stabilized by H-bonds, but without proximity
• Figure 6.3b, page 165
          Types of Beta Sheets
• Parallel- adjacent segments proceed in same
  directions as far as amino to carboxyl
• Antiparallel- Adjacent chains proceed in
  opposite directions, as in 6.3b.

• Most have a right hand twist, they are not flat

• Depictions, see figure 6.16, page 179
            Loops and Bends
• Approximately half the residues in a protein
  are in either a helix or beta sheet.
• The other half are in less ordered
  conformations such as Loops, Bends, etc.
• Turns and Bends are short segments that
  connect secondary structures
                  Beta Turn
• Beta turns involve four Amino Acid residues
  with the 1st residue H-bonded to the 4th
  residue resulting in a tight, 180 degree turn

• Fig 6.18 page 181

• Proline and Glycine are often present in beta
  turns. Why?
• Loops are regions that contain more AA
  residues than minimally required to connect
  adjacent secondary structures
• Although they have no set conformation, they
  do serve key biological roles
• There are some conformations that are
  secondary structures interacting, but not
  enough to be termed a tertiary
     Super Secondary Structures
• Helix-Loop-Helix
  – Provide the oligonucleotide-binding portion of
    DNA-binding proteins such as repressors and
    transcription factors
                Back to Loops
• Loops are mainly found on the exterior
  portion of a protein.
  – Exposed to solvent
  – Readily accessible sites for recognition and
• While the structure may be irregular, they still
  exist in specific conformations
• Conformations are held together by H-bonds,
  salt bridges, and hydrophobic interactions
 Tertiary and Quaternary Structure
• 3o refers to the entire 3D conformation of the
• Domain-a section of protein structure
  sufficient to perform a particular chemical or
  physical task such as binding of a substrate or
• Proteins may have one or more domains.
• Fig 6.27 page 192
 Tertiary and Quaternary Structure
• In some cases, proteins are assembled from
  more than one polypeptide or protomer
• Quaternary structure defines both the
  composition and spatial relations between
  these multiple chains.
• Monomeric proteins consists of one chain
• Dimeric proteins consists of two chains
• For Dimeric and above, there are different
  – Homodimers-contain two copies of same chain
  – Heterodimers- contain two different chains
• Greek letters are used to identify the different
  chains and subscripts indicate the number of
  each unit.
     Stability of 3o and 4o structures
•   Primarily stabilized by non-covalent forces
•   Hydrophobic interactions-
•   H-Bonds
•   Salt-bridges
•   Etc

• Think Velcro!!!
      Determining 3D Structure
• Two main techniques:
• X-Ray crystallography

• NMR Spectroscopy
         X-Ray Chrystallography
• Key is to get a good crystal!!!

• Then bombard the crystal with x-rays and
  acquire a diffraction pattern

• The diffraction pattern, along with the primary
  structure can be used to deduce the 3D
           X-Ray Chrystallography
• Computers are getting better and better at
  interpreting X-ray diffraction patterns
• Major stumbling block remains the requirement of
  inducing highly purified samples of the protein to
• Several lines of evidence, including the ability of
  some crystallized enzymes to catalyze chemical
  reactions, indicate that the majority of the
  structures determined by X-ray crystallography
  represent the structures of proteins in free solution.
           NMR Spectroscopy
• Many common nuclei can be “seen” by NMR

• Chemical shifts depend on the nuclei, what
  functional group it is in, and neighboring NMR
                 2-D NMR
• Shows which nuclei are near each other.
             Protein Folding
• We have said the number of possible
  conformations of even small peptides is very
• Thermodynamics dictate the formation of the
• The desired conformation is usually
  energetically favored
                 Protein Folding
• Even with this energetically favored situation, it
  would take billions of years for a protein to explore
  all possible conformations to find the favored state
• So folding has to take place in some sort of guided
• First, as protein leaves the ribosome, short
  segments fold into secondary structures which are
  directed by the primary sequence
              Protein Folding
• Second, the aqueous environment of the cell
  drives all hydrophobic side groups to the
  middle of the protein creating a molten
• Regions of secondary structure rearrange to
  form the mature conformation.
• While this process is ordered, it is not rigid
• In vivo- in the cell
• In vitro- in test tube
• Denatured- unfolding of a protein

• Aggregates- disordered complexes of unfolded
  or partially unfolded polypeptides held
  together by hydrophobic interactions
• Chaperone proteins help fold over half the
  proteins in mammals
• They work by covering up hydrophobic
  groups, shielding them from the aqueous
  environment, thus preventing aggregation
• Chaperons can also rescue misfolded proteins.
              Higher Orders
• Disulfide bonds help stabilize tertiary and
  quaternary structures even though they are
• The protein disulfide isomerase is present to
  continually break and reform disulfide bonds.
       X-Proline Peptide Bonds
• All X-proline peptide bonds-where X
  represents any residue-are synthesized in the
  trans configuration.
• However, of the X-Proline bonds of mature
  proteins, approximately 6% are cis.
• The cis configuration is particularly common in
• Isomerization from trans to cis is catalyzed by
  the enzyme proline-cis,trans-isomerase.
           Alzheimer’s Disease
• Refolding or misfolding of protein endogenous
  to human brain tissue, b-amyloid, is a
  prominent feature of Alzheimer’s Disease.
• Levels of b-amyloid become elevated, and this
  protein undergoes a conformational
  transformation from a soluble helix-rich state
  to a state rich in beta sheets and prone to self-
           Mass Spectrometry
• Separates molecules based on MW
• Peptides are vaporized
• Applied charge propels the molecules down a
  bent tube
• Molecules interact with magnetic field in tube
• Where the molecules hit the side of the tube,
  which is the detector, gives their mass
           Mass Spectrometry
• Since all A.A. except Leu and Isoleu have
  different masses, all AA present can be

• Post translational modifications can also be
  determined with this method
Posttranslational Modifications
  Modification     Mass Increase (Da)
 Phosphorylation          80
  Hydroxylation           16
  Methylation             14
   Acetylation            42
  Myristylation           210
 Palmitoylation           238
  Glycosylation           162
• The methods of vaporization vary and recent
  advances have allowed for larger peptides to
  be used.

• Book discusses “Time of Flight” MS.
         Make sure you read!!
• Make sure you read Ch. 6. Remember, you are
  responsible for material not covered in
• Example:
• Fibrous Proteins: Structural materials of Cells
  and tissue. Be able to compare and contrast
  Karatins, Fibron, collagen, and elastin.

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