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Membrane Partitioning/Membrane Binding
Binding to Proteins:
•Specific well defined binding sites
•Number of binding sites per molecule
•Strength of the interaction
•cooperativity
Adsorption of Ions and Amphipathic molecules to bilayers is not always as
straightforward – binding site may not be as well defined. There may be
specific ligands, there may be hydrophobic partitioning, or electrostatic
attraction
Different Models of Binding:
Partition Equilibrium
Langmuir Adsorption Isotherms
Complexation to N Lipids
What do we mean by
BINDS?
Partition Equilibrium
P + L  PL
P = small molecule (protein etc)
L = lipid, lipid is considered a separate phase
Partitioning Coefficient: Kp = Cb/Cf
Where Cb is the concentration of bound molecule (PL), Cf is the
concentration of free molecule in solution (P).
Binding Isotherms are typically analyzed by measuring either the amount of
the free ligand in solution or that bound to the bilayer and knowing the total
concentration of lipid.
An expression was given in Fridays paper presentation:
Typically you derive an expression in terms of known total amounts, measure
one parameter to determine a binding constant
Langmuir Adsoprtion Isotherm: membrane is treated as a lattice of potential
binding sites – often not relevant to membrane binding phenomenon
Complexation to n-lipids
L + nP
LPn
L = ligand, small molecule
P = phospholipid
Ka = [LPn] / ([L][P]n) --- resulting equations are referred to as Scatchard Plots
Specific Ligand Interactions, Electrostatics, Dehydration all play a role in binding
Classes of Ligands Which Interact with the Lipid Bilayer
•Class I :Non-Polar Solutes
•Class II: Amphipathic Molecules
•Anesthetics •Drugs – antipsychotics, antianxiety etc - chlorpromazine
•Antibiotics
•Detergents
•Membrane Probes
•Class III: Hydrophobic Ions
•Class IV: Ions
Nonpolar molecules
Class I: Benzene, hydrocarbons,
perfloroalkanes
PFOB
Antimicrobial Peptides Antibiotics/Antifungals
Mode of interaction is based upon specific properties of the peptide and target
membrane
Many microbial antibiotics are peptides that form cationic amphipathic secondary
structures that interact with negatively charged bacterial membranes via aid of
electrostatic interactions. – form pores, leading to membrane permeabilization
•Amphipathic/hydrophobic a-helices-magainin
b-sheet peptides and small proteins-defensins
•Peptides with irregular AA composition•Peptides with thio-ether rings-lantibiotics
•Peptaibols –alamethicin Aib
•Macrocyclic cysteine knot peptides
Lytic peptides: eukaryotic, prokaryotic, both
BBA 1999 1462, issues 1-2
Peptide-Membrane Interactions:
membrane-peptide mediated : can change sterochemistry and still have lytic
effects, in fact, gramicidin is composed of altering L and D amino acids
Most anit bacterial peptides contain high charge and amphipathic nature
Hemolytic peptides : low net positive or negative charge.
Bee Venom : Mellitin
Frog toxin: Meganin
Antifungals: Nystatin and Amphotericin B (polyene macrolide)
Fungal and mammallian cells – recognize sterols, more strongly ergosterol
than cholesterol (basis for fungal selectivity)
•Only in contact with the head groups
•Secondary structure unimportant
•Not inserted into hydrophobic core
•Monomers bind in a-helical state
•Monomers recognize and assemble
•Helices insert into hydrophobic core
•Additional recruitment of monomers
Detergent interactions with Bilayers: last week lecture and presentation.
Small Probe Molecule: EPR and Fluorescent Molecules
Membrane “Fluidity” Measured by TEMPO partitioning
f = fraction in the bilayer
Hyperfine splitting is sensitive to polarity
H is “bilayer”
P is aqueous buffer
Shows Gel to liquid transition
Bilayer Permeability: (a) enter membrane, (b)
diffuse across (c)exit membrane
P = KpDm/d
Kp = Cm/Caq