Detergents amphipathicamphiphillic molecules that have a limited solubility and above a critical con

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Detergents amphipathic/amphiphillic molecules
that have a limited solubility and above a
critical concentration form micelles (CMC).
If detergent concentration is less than CMC,
monomers are in solution
If detergent concentration is above CMC, ..
When micelles are present, there is always free
monomer present too
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Detergents are typically broken down into 4
classes
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General Properties of Classes of Detergents for
Membrane Protein Studies
SDS will help stabilize an all a-helix
peptide/protein.
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Solubilizing Membranes
Protein Purification Schemes
Membrane Protein Reconstitution

• Detergent Elimination Strategies
• Dialysis
• Dilution
• Size Exclusion
• Biobeads (hydrophobic adsorption)

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Features of the 4 stages...
Ratio of detergent/lipid important, detergent
important
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Turbidity and Light Scattering
An increase in turbidity/light scattering is an
indication of a larger size particle (as long as
the wavelength used is larger than the particle
size). The decrease is taken as
solubilization. Turbidity is often measured in
simple spectrophotometer ( 300-500nm) At
solubilizing concentrations, most detergents will
actually increase turbidity shown to result from
reassembly into multilammellar structures at
point of solubilization.
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Turbidity and Light Scattering
light scattering performed as 90o scattered
light under steady state conditions using a
spectrofluorometer. It is more sensitive than
turbidity, can have smaller sample sizes and go
to longer wavelengths. dynamic light
scattering quasielastic light
scatteringphoton correlation spectroscopy the
intensity of scattered light is monitored in the
ms time range. The Brownian motion of the
particles induces a broadening of the spectrum,
in a way that is related to their size and
shape. You can determine round from disk from
tubes etc.
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Entire Issue of BBA biomembranes Volume 1508,
Nov 2000
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Effects of Salt and Urea
Vesicle solubilization
Salt Effects on detergent CMC
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Studies go both ways Solubilization of Liposomes
by Detergents Removal of Detergents to Form
Proteoliposomes.
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