Protein Structure and Function

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Chapter 4

• Protein Structureand Function

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Proteins

• Make up about 15 of the cell
• Have many functions in the cell
• Enzymes
• Structural
• Transport
• Motor
• Storage
• Signaling
• Receptors
• Gene regulation
• Special functions

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Shape Amino Acid Sequence

• Proteins are made of 20 amino acids linked by
  peptide bonds making a polypeptide
• Polypeptide backbone is the repeating sequence of
  the N-C-C-N-C-C following the peptide bond
• The side chain or R group is not part of the
  backbone or the peptide bond

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Polypeptide Backbone
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Amino AcidsNOTE You need to know this table
Hydrophilic
Hydrophobic
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Protein Folding

• The peptide bond allows for rotation around it
  and therefore the protein can fold and orient the
  R groups in favorable positions
• Weak non-covalent interactions will hold the
  protein in its functional shape these are weak
  and will take many to hold the shape

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Non-covalent Bonds in Proteins
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Globular Proteins

• The side chains will help determine the
  conformation in an aqueous solution

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Hydrogen Bonds in Proteins

• H-bonds form between 1) atoms involved in the
  peptide bond 2) peptide bond atoms and R groups
  3) R groups

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Protein Folding

• Proteins shape is determined by the sequence of
  the amino acids
• The final shape is called the conformation and
  has the lowest free energy possible
• Denaturation is the process of unfolding the
  protein
• Can be down with heat, pH or chemical compounds
• In the chemical compound, can remove and have the
  protein renature or refold

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Refolding

• Molecular chaperones are small proteins that help
  guide the folding and can help keep the new
  protein from associating with the wrong partner

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Prion Proteins
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Shape Dictates Function
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Protein Folding

• 2 regular folding patterns have been identified
  formed between the bonds of the peptide backbone
  (N-H and CO)
• ?-helix protein turns like a spiral fibrous
  proteins (hair, nails, horns)
• ?-sheet protein folds back on itself as in a
  ribbon globular protein

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? Sheets

• Core of many proteins is the ? sheet
• Form rigid structures with the H-bond
• Can be of 2 types
• Anti-parallel run in an opposite direction of
  its neighbor (A)
• Parallel run in the same direction with longer
  looping sections between them (B)

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? Helix

• Formed by a H-bond between every 4th peptide bond
  CO to N-H
• Usually in proteins that span a membrane
• The ? helix can either coil to the right or the
  left
• Can also coil around each other coiled-coil
  shape a framework for structural proteins such
  as nails and skin

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CD from Text

• The CD that is included on your textbook back
  cover has some video clips that will show the ?
  helix and ? sheets as well as other things in
  this chapter. You will want to look at them. If
  you have problems, we will look at them during
  lab.

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Levels of Organization

• Primary structure
• Amino acid sequence of the protein
• Secondary structure
• H bonds in the peptide chain backbone
• ?-helix and ?-sheets
• Tertiary structure
• Non-covalent interactions between the R groups
  within the protein
• Quanternary structure
• Interaction between 2 polypeptide chains

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Domains

• A domain is a basic structural unit of a protein
  structure distinct from those that make up the
  conformations (100-250 amino acids)
• Part of protein that can fold into a stable
  structure independently
• Different domains can impart different functions
  to proteins
• Proteins can have one to many domains depending
  on protein size

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Protein Structure
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Domains
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Useful Proteins

• There are thousands and thousands of different
  combinations of amino acids that can make up
  proteins and that would increase if each one had
  multiple shapes
• Proteins usually have only one useful
  conformation because otherwise it would not be
  efficient use of the energy available to the
  system
• Natural selection has eliminated proteins that do
  not perform a specific function in the cell

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Protein Families

• Have similarities in amino acid sequence and 3-D
  structure
• Have similar functions such as breakdown proteins
  but do it differently

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Proteins Multiple Peptides

• Non-covalent bonds can form interactions between
  individual polypeptide chains
• Binding site where proteins interact with one
  another non-covalent bonds
• Subunit each polypeptide chain of large protein
• Dimer protein made of 2 subunits
• Can be same subunit or different subunits

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Single Subunit Proteins
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Different Subunit Proteins

• Hemoglobin
• 2 ? globin subunits
• 2 ? globin subunits

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Protein Assemblies

• Proteins can form very large assemblies
• Can form long chains if the protein has 2 binding
  sites link together as a helix or a ring
• Actin fibers in muscles and cytoskeleton is
  made from thousands of actin molecules as a
  helical fiber

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Shapes
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Types of Proteins

• Globular Proteins most of what we have dealt
  with so far
• Compact shape like a ball with irregular surfaces
• Enzymes are globular
• Fibrous Proteins usually span a long distance
  in the cell
• 3-D structure is usually long and rod shaped

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Important Fibrous Proteins

• Intermediate filaments of the cytoskeleton
• Structural scaffold inside the cell
• Keratin in hair, horns and nails
• Extracellular matrix
• Bind cells together to make tissues
• Secreted from cells and assemble in long fibers
• Collagen fiber with a glycine every third amino
  acid in the protein
• Elastin unstructured fibers that gives tissue
  an elastic characteristic

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Collagen and Elastin
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Stabilizing Cross-Links

• Disulfide bonds (S-S) form between adjacent -SH
  groups on the amino acid cysteine
• Cross linkages can be between 2 Cys of a single
  protein or between 2 subunits making up the
  protein
• S-S bond is made in the ER as it is necessary for
  extracellular proteins and not for cytosolic
  proteins

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Proteins at Work

• The conformation of a protein gives it a unique
  function
• To work proteins must interact with other
  molecules, usually 1 protein with 1 or a few
  molecules from the thousands in cell
• Ligand the molecule that a protein can bind
• Binding site part of the protein that interacts
  with the ligand
• Consists of a cavity formed by a specific
  arrangement of amino acids

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Ligand Binding
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Formation of Binding Site

• The binding site forms when amino acids from
  within the protein come together in the folding
• Amino acids in binding site come from throughout
  the primary structure
• The remaining sequences may play a role in
  regulating the proteins activity

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Antibody Family

• A family of proteins that can be created to bind
  to almost any molecule
• Antibodies (immunoglobulins) are made in response
  to a foreign molecule ie. bacteria, virus,
  pollen called the antigen
• Bind together tightly and therefore inactivates
  the antigen or marks it for destruction

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Antibodies

• Y-shaped molecules with 2 binding sites at the
  upper ends of the Y
• The loops of polypeptides on the end of the
  binding site are what imparts the recognition of
  the antigen
• Changes in the sequence of the loops make the
  antibody recognize different antigens -
  specificity

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Antibodies
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Enzymes as Catalysts

• Enzymes are proteins that bind to their ligand as
  the 1st step in a process
• An enzymes ligand is called a substrate
• May be 1 or more molecules
• Output of the reaction is called the product
• Enzymes can repeat these steps many times and
  rapidly, called catalysts
• Many different kinds see table 4-1, p 147

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Enzymes at Work

• Lysozyme is an important enzyme that protects us
  from bacteria by making holes in the bacterial
  cell wall and causing it to break
• Lysozyme adds H2O to the glycosidic bond in the
  cell wall
• Lysozyme holds the polysaccharide in a position
  that allows the H2O to break the bond this is
  the transition state state between substrate
  and product
• Active site is a special binding site in enzymes
  where the chemical reaction takes place

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Lysozyme

• Non-covalent bonds hold the polysaccharide in the
  active site until the reaction occurs

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Features of Enzyme Catalysis
Proper orientation
Alter e- distribution
Change shape
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Prosthetic Groups

• Occasionally the sequence of the protein is not
  enough for the function of the protein
• Some proteins require a non-protein molecule to
  enhance the performance of the protein
• Hemoglobin requires heme (iron containing
  compound) to carry the O2
• When a prosthetic group is required by an enzyme
  it is called a co-enzyme
• Usually a metal or vitamin
• These groups may be covalently or non-covalently
  linked to the protein

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Regulation of Enzymes

• Regulation of enzymatic pathways prevent the
  deletion of substrate
• Regulation happens at the level of the enzyme in
  a pathway
• Feedback inhibition is when the end product
  regulates the enzyme early in the pathway

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Feedback Regulation

• Negative feedback pathway is inhibited by
  accumulation of final product prevents enzyme
  from working
• Positive feedback a regulatory molecule
  stimulates the activity of the enzyme, usually
  between 2 pathways
• ? ADP levels cause the activation of the
  glycolysis pathway to make more ATP

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Allostery

• Conformational coupling of 2 widely separated
  binding sites must be responsible for regulation
  active site recognizes substrate and 2nd site
  recognizes the regulatory molecule
• Protein regulated this way undergoes allosteric
  transition or a conformational change
• Protein regulated in this manner is an allosteric
  protein

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Allosteric Regulation

• Method of regulation is also used in other
  proteins besides enzymes
• Receptors, structural and motor proteins

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Allosteric Regulation

• Enzyme is only partially active with sugar only
  but much more active with sugar and ADP present

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Phosphorylation

• Some proteins are regulated by the addition of a
  PO4 group that allows for the attraction of
  charged side chains causing a conformation change
• Reversible protein phosphorylations regulate many
  eukaryotic cell functions turning things on and
  off
• Protein kinases add the PO4 and protein
  phosphatase remove them

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Phosphorylation/Dephosphorylation

• Kinases capable of putting the PO4 on 3 different
  amino acid residues
• Have a OH group on R group
• Serine
• Threonine
• Tyrosine
• Phosphatases that remove the PO4 may be specific
  for 1 or 2 reactions or many be non-specific

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GTP-Binding Proteins (GTPases)

• GTP does not release its PO4 group but rather the
  guanine part binds tightly to the protein and the
  protein is active
• Hydrolysis of the GTP to GDP (by the protein
  itself) and now the protein is inactive
• Also a family of proteins usually involved in
  cell signaling switching proteins on and off

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Molecular Switches
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Motor Proteins

• Proteins can move in the cell, say up and down a
  DNA strand but with very little uniformity
• Adding ligands to change the conformation is not
  enough to regulate this process
• The hydrolysis of ATP can direct the the
  movement as well as make it unidirectional
• The motor proteins that move things along the
  actin filaments or myosin

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Protein Machines

• Complexes of 10 or more proteins that work
  together such as DNA replication, RNA or protein
  synthesis, trans-membrane signaling etc.
• Usually driven by ATP or GTP hydrolysis
• See video clip on CD in book