Transcript LECT12 allo2aB

Interpreting Effects of Allosteric Agents
100
Bind O2
95
Fractional
Saturation
50
Shift to
R state
80
O2
OH-Hb
10
pO2
Shift to
T state
CO2
BPG
H+
Release O2
4 RULES Governing Allosteric Proteins
RULE: Allostery is a property of proteins that contain
multiple subunits or a single subunit with multiple binding
sites.
RULE: Allosteric proteins must be able to bind more than
one ligand.
RULE: A ligand has the capacity to induce a change in
the shape of an allosteric protein.
RULE: By changing the protein’s shape, a ligand can
facilitate (positive) or deter (negative) the binding of
additional ligands to the protein.
MODELS OF ALLOSTERY
CONCERTED: (ALL OR NONE)
See Strategies
p.120
Subunits exist in two conformational states
The two states are in equilibrium
All subunits have the same conformation; no hybrids
Ligand binds to any subunit in either conformation
Conservation of symmetry is the driving force
Problems with Concerted Model
• Symmetry is not preserved in most
oligomeric proteins
• Some allosteric oligomeric proteins do not
have identical subunits
• Cannot explain in a two state model how
negative and positive cooperativity occur in
the same protein
Sequential Model
S
S
S
S
S
S
S
S
S
S S
S
Ligand binding induces change in subunit
Change is progressive
Symmetry is not preserved
Mechanical coupling between subunits may be weak
S
S
Sickle Cell Anemia
(a genetic disease)
One Amino acid in a
protein can make a
difference between
life and death
Distortion
Is it a membrane protein?
Is it a cytosolic protein?
Is it something other than a protein?
Hb (alpha) N- Val-Leu-Ser-Pro-Ala-Asp-Lys-Thr……..
Hb (beta) N- Val-His-Leu-Thr-Pro- Glu
Val-Glu-Lys…….
Sickle Cell
Glu6  Val6 (each beta chain)
Glu
GAA or GAG
Val
GUA or GUG
Transposition
A U replaces an A in the codon (a pyrimidine/purine exchange)
Val
Polymerized deoxy Hb Aggregates 
Insoluble fibers
Distorts the overall shape of hemoglobin
Distorts the overall shape of an erythrocyte
Distorted erythrocytes cannot pass through capillaries
Antibodies
Ancient Greeks:
Thucydides (460-400 B.C.) Speaking of the plague of Athens,
Yet, it was in those that had recovered from the
disease that the sick and dying found most
compassion. These knew what it was from experience,
and had now no fear for themselves; for the same
man was never attacked twice-never at least fatally.
Early Theories
Pauling 1940: The “Instructive” Theory
Antigens act like templates that direct
the folding of a nascent antibody chain
MacFarlane Burnet: The Selective or clonal theory
The combining site on an antibody molecule
is completely determined before it encounters
an antigen
Bone Marrow
Precursor
lymphocytes
leukocytes
macrophages
B-lymphocytes
T-lymphocytes (T-cells)
Antibodies
Cytotoxic T cells
(TC cells)
Cell-Mediated
Helper T cells
(TH cells)
Cytokines
Humoral
Essential Features
• Each antibody-producing B-cell makes a single
kind of antibody…no antigen is needed.
• Specificity is determined by amino acid
sequence…each cell’s DNA is distinctive.
• Immature cells that make antibodies are
destroyed early in life…self tolerance
• Mature cells make and display antibodies on
their surface
• Interaction with antigen triggers cell to divide,
making large amounts of a particular antibody
that persists until antigen is gone
Antibodies are large complex molecules
Composition:
Antigen-binding site
Constant
Variable
Hinge region
2 heavy, 2 light chains
Disulfide bonds
5 Classes of Antibodies (Immunoglobulins, Ig)
• IgA…external secretions, tears,saliva,
bronchial and intestinal mucous
• IgG…principal antibody in the serum,
originally called gamma globulin
• IgD…least understood
• IgE…no clear function, has receptor on
mast cells and stimulates production of
histamine and is linked to allergic responses
• IgM…first class to appear, highest
combining sites, effective against bacteria
Mechanism
• Preexisting B cell synthesizes IgM first
• IgG, IgA, IgD, IgE of same specificity are
made later
• Light chain is unchanged during switch
• Variable region of heavy chain is unchanged
during switch
• Only the Constant region of heavy chain
changes…(class switching or CH switching)
• In the mouse, switching uses the appropriate
Constant region gene, [C, C, C, C, C] for
the antibody class
Cell-Mediated Immunity
Major histocompatibility complex (MHC)
Function: In conjunction with T-cell surveillance,
MHC is a series of polymorphic proteins designed to
display digested peptide fragments on the surface of
the cell. The basis of cell-mediated immunity
MHC-1
On all cell surfaces
Displays fragments from internal protein digests
Interacts with TC cells to initiate infected cell destruction
MHC-2
On surface of phagocytic cells (macrophages, etc.)
Displays digested fragments from external protein sources
Interacts with TH cells to alert system of infection
Important!
Internal proteins digested by proteases cannot bind to
MHC-2
MHC-1 interaction is a signal to TC cells to
destroy the infected cell
External proteins that enter the cell by an endosomal
pathway cannot bind to MHC-1.
MHC-2 interaction is a signal to TH cells
that the system is under attack by a virus,
bacteria, etc. and to send for help in the
form of antibody-producing cells