Download Chem331 Lect 14 Membranes

Membranes
What are the purposes of membranes?
Physical barriers/compartmentalization
Gatekeepers—exclusion of toxic molecules
Energy and signal transduction
Aid in cell locomotion
Cell-cell interactions
Lipid and Protein Concentration Vary in
Different Types of Membranes
Depending on the function of the membrane
(structural vs. a place where reactions are
catalyzed), the protein and lipid content
changes
Membranes that carry out enzyme reactions,
signal transduction, transport, etc. are typically protein rich
Lipid composition also varies
Lipids Order in Aqueous Solutions
Amphipathic lipid form a variety of structures in water RAPIDLY.
Micelles—lipid polar head faces outward, hydrophobic tails are
inward. Typically made of a few hundred molecules.
Critical Micelle Concentration: concentration of lipids needed to
form micelles. Below CMC—individual lipids
Ordered Structures of Lipids - Bilayers form spontaneously
over large areas
Bilayers minimize solvent exposure by wrapping around
themselves—lamellar structures
Membrane Thickness and Shape
Typical cell membranes are approximately 50Å thick (includes the lipid bilayer and membrane
proteins)
Thickness is defined by particular lipid/protein composition of the membrane
–
Some membrane proteins stick far out of the lipid bilayer and can change thickness
by as much as 5Å!
Though we draw lipids tails as being perpendicular to the membrane plane, they can actually
tilt/bend (think about all those unsaturated fatty acids and their particular conformations!)
Fluid Mosaic Model
Discovered by SJ Singer and GL Nicolson in 1972
Membranes are dynamic structures composed of proteins and phospholipids
Phospholipid bilayer is a fluid matrix—2D solvent for proteins.
2 types of proteins in bilayer:
–
Peripheral/Extrinsic proteins
• don’t penetrate bilayer
• associated by non-covalent interactions to surface of bilayer or embedded
proteins
• dissociate by changing salt concentration/pH
–
Integral/Intrinsic proteins
• have hydrophobic surfaces that penetrate bilayer and regions that interact
with the aqueous environment
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Cholesterol Impacts Membrane Fluidity—Prevents Extremes
-OH group on cholesterol interacts with polar head groups,
steroid/hydrocarbon chain buried in the lipid bilayer
Decreases membrane fluidity, increases membrane packing—also
prevents membrane crystallizaton
Reduces the membrane’s permeability to neutral solutes, protons, and
other ions—a good thing!
Membranes are More Mosaic than Fluid!
Peripheral MembraneProteins
LOOSELY associated with membrane via 4 different types of interactions:
Integral Membrane Proteins: Single Transmembrane Segments
Proteins anchored in membrane by a single hydrophobic segment—typically an α-helix.
10-30% of transmembrane proteins have a single helical transmembrane segment
Typically function as cell surface receptors for extracellular
signaling or immune recognition sites
Example: Glycophorin A Spans red blood cell membrane
Glycoprotein—oligosaccharides dictate MN antigenic
specificities
Integral Membrane Proteins: Multiple Transmembrane
Segments
Cross the lipid bilayer more than once
Typically have 2-12 transmembrane segments
Example: Bacteriorhodopsin
•
light driven proton transport for
Halobacterium
•
Primary sequence: 7 segments
about 20 nonpolar residues in
length
Hydropathy Plots - Give us clues to
determine whether or not a polypeptide
sequence with a given hydrophobicity
would likely be included in a membrane
Beyond the α-helix: β-strands span the membrane too!
β-barrels
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–
–
–
Maximize H-bonding, very stable
Interior can accommodate H2O molecules and peptide chains
Polar and non-polar residues alternate along the strands, polar face in (to interact w/
water), nonpolar face out and interact with lipids
β-barrel Examples
Porins: Maltoporin
–
transports maltose through E. coli outermembrane
Multiple Subunit β-barrel: α-hemolysin toxin (7-mer)
–
secreted as monomers by Staph. aureus
–
Channel allows uncontrolled permeation of water, ions, small molecules
Membranes are Both Heterogeneous and Asymmetric
Lateral heterogeneity—certain types of proteins and lipids cluster together in the membrane
plane
Transverse asymmetry—the inner and outer leaflet of a membrane may have different protein
and lipid compositions
Example: Typical animal cell—amine-containing phospholipids enriched in the cytoplasmic
(inner) leaflet of the plasma membrane, and choline containing phospholipids and sphingolipids
enriched in outer leaflet
Imbalances in these levels and loss of transverse asymmetry can result in catastrophe—cell
death signaling
Moving Lipids Modulate Membrane Functions
Lateral diffusion of lipids can occur quickly (could move in a linear direction several microns per
sec.)
Proteins can help move lipids from one side of the bilayer to the other
–
Flippases—ATP dependent, specific lipids
–
Floppases—ATP dependent, specific lipids
–
2+
Scramblases—ATP independent, non-specific, requires Ca
Flippases, Floppases and Scramblases Help Maintain Transverse Asymmetry
Membrane Ordering
Membrane Ordering
Low Temps—gel phase/solid-ordered state, acyl chain perpendicular to membrane plane, chains
packed, very little movement
Higher Temps— liquid crystalline/liquid-disordered state, acyl chains in motion-rotation around
C-C bonds and increased disorder
Moving between these two states is
called a phase transition, and
occurs at a transition
temperature/melting temperature
Sharpness of transition indicative of
cooperative behavior—molecules in
same vicinity acting in concert
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Membrane Rafts
The liquid ordered state—acyl chain ordering, but translational disorder
Up to 50% of the plasma membrane consists of these rafts
Small and transient—hard to measure!
Hopping the Fence
Kusumi et al. determined using single particle tracking of fluorescently labeled lipids that lipids
tend to stay in compartments, but occasionally “hop over the fence” to new compartments.
Transport Across Membranes
Passive Diffusion
–
Uncharged Molecule: Entropic process, movement across
membrane happens until concentration of molecules the
same on either side
–
Charged Molecule: Dependent on electrochemical potential
Transport Across Membranes
FACILITATED DIFFUSION: Passive
diffusion is too slow to sustain most
biochemical processes
Facilitated diffusion occurs via proteins
with net movement of solvent happening in
a thermodynamically favored direction
(dG < 0)
These proteins typically have a high binding affinity for certain
solutes
Membrane Channel Proteins
Single Channel Pores: Made up of dimer, trimer, etc. multimeric protein subunits :
Multimeric subunit assemblies where each subunit has its own pore:
Facilitated Diffusion Membrane Channel Proteins
Channels are often selective for a particular type of ion or molecule.
–
Usually have their pores lined with amino acids of the opposite charge of the ion
they are transporting
Some are gated—open/close upon a signal.
–
Voltage-gated respond w/ change in membrane potential.
–
Ligand gated open/close with binding of a specific ion, molecule, small protein
Depending on channel size, ions can flow through in a hydrated or dehydrated state
–
Wide, non-selective
–
Narrow, charged amino acids lining, highly selective
Example: Potassium Channels
Potassium transport is critical for maintaining cell volume, electrical impulse formation (neurons)
K+ channels are highly selective for K+ over Na+, and conduct K+ ions at very fast rates—at the
diffusion limits
For the tetramer channel KcsA, four pentapeptides ThrValGlyTyrGly (backbone carbonyls and
Thr oxygens) mediate selectivity of K+ vs Na+
–
Selectivity based on atomic radius!
Facilitated Transport: Potassium Channels
High selectivity and quick transport—seems
paradoxical!
Repulsion from closely spaced K+ ions and
conformational changes induced by binding keep things
moving….
Channel Conformational Changes Leave the
Channel Open or Closed
pH induced helix bending and rearrangement 4
Active Transport
Active Transport—transport of species from low to high concentrations—requires energy
input
–
Energy sources come from ATP hydrolysis, light energy and energy stored in ion
gradients.
Monvalent cation transport coupled to ATP-hydrolysis
+ +
–
Na ,K -ATPase—sodium pump
+
+
Na ,K -­‐ATPase
Integral membrane protein with 3 subunits
–
alpha (120 kDa), beta (35 kDa),
gamma (6.5 kDa)
Actively pumps 3 Na+ out of cell and 2 K+ into
cell for every one ATP hydrolyzed
+
+
Na ,K -ATPase Inhibitors
Cardiac glycosides/Cardiotonic steroids
–
Bind to extracellular side of
+ +
Na ,K -ATPase to form very stable complex
People with high blood pressure/hypertension have high levels of these inhibitors.
Accumulation of Na+ and Ca2+ narrows the vessels and creates hypertension
ABC Transporters
ATP-Binding Cassette (ABC) transporters. Superfamily of transporters found across most ancient
and modern cells.
- Active transporters which use ATP to drive compound transport against gradient.
- Transport ions, lipids, foreign compounds even nucleotides and membranes
ABC transporters (over 1,100 different ABC transporters across all organisms / 50 in humans)
– 2 transmembrane domains, form a pore; cytosolic nucleotide binding domains
– Conserved core of 12
transmembrane helices
- Mechanism of transport changes
depending on specific transporter
- 15 different known defects altering 14
of the 50 human transporters lead to
known diseases
Multidrug resistance (MDR) efflux
pumps (over expressed in some tumor
cells)
- export cellular waste molecules and
toxins
- can be a problem for certain
therapeutic drugs
- Other diseases associated with ABC transporters include: Cystic fibroses, age related
macrodegeneration, and others
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