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The Three Dimensional Structure of Proteins

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					           Chapter 6
The Three-Dimensional Structure
          of Proteins
           Homework:
            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
  conformations
• 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
  residues
• 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
• 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
    binding
• 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
  protein
• Domain-a section of protein structure
  sufficient to perform a particular chemical or
  physical task such as binding of a substrate or
  ligand
• 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.
               Designations
• Monomeric proteins consists of one chain
• Dimeric proteins consists of two chains
• For Dimeric and above, there are different
  kinds:
  – Homodimers-contain two copies of same chain
  – Heterodimers- contain two different chains
                Designations
• 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
  structure.
           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
  crystallize
• 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
  nuclei
                 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
  large
• Thermodynamics dictate the formation of the
  conformation
• 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
  environment
• 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
  globule
• Regions of secondary structure rearrange to
  form the mature conformation.
• While this process is ordered, it is not rigid
                Definitions
• 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
               Chaperones
• 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
  non-specific.
• 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
  Beta-turns.
• 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-
  aggregaton.
           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
  determined

• 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
                 Advances
• 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
  lecture.
• Example:
• Fibrous Proteins: Structural materials of Cells
  and tissue. Be able to compare and contrast
  Karatins, Fibron, collagen, and elastin.

				
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posted:9/1/2011
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