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Membrane Protein Folding and Stability

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What are the dominant forces that determine structure and stability? ... Predicting of structure and stability from sequence is fundamentally a problem ... – PowerPoint PPT presentation

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Title: Membrane Protein Folding and Stability


1
Membrane Protein Folding and Stability
  • Physical Principles

Episode 1 Introduction, Partitioning into
membranes
http//www.biochem.oulu.fi/juffer/
2
Membrane proteins (MPs) (1)
  • What are the general features of MPs?
  • How are they assembled?
  • How can their thermodynamic stability be
    described?
  • What is the nature of the lipid bilayer as a host
    phase for MPs?
  • What are the dominant forces that determine
    structure and stability?
  • What can the folding and stability of MPs tell
    us about the protein folding problem generally?

3
Membrane proteins (MPs) (2)
  • Two structural motifs
  • a-helix bundles
  • b-barrels

1.5 nm
3 nm
b-barrels
a-helix bundle
Trp, Tyr side chains located typically in
interfacial region.
HChydrocarbon
4
How many MP structures?
5
MP versus soluble proteins
  • MP are generally not soluble in water.
  • MP interior is similar to those of soluble
    proteins.
  • MP interior comprised of internally H-bonded
    a-helices and b-sheets.
  • MP have direction in space
  • Lys, Arg are more abundant in cytoplasmic domain
    relative to periplasmic domain.
  • Tyr, Trp display strong preference for
    interfacial region.

6
Assembly of MPs (1)
  • Translocation apparatus involving the transient
    attachment of a active ribosome to a translocon
    embedded in membrane.
  • Spontaneous insertion of soluble protein into
    membrane.

7
Assembly of MPs (2)
  • Regardless of insertion/assembly process
  • Stably folded MPs reside in a free energy
    minimum.
  • Minimum is determined by net energies of the
    interaction of peptide chains with
  • Water.
  • Each other.
  • Lipid bilayer (hydrocarbon core and interface).
  • Co-factors.
  • Predicting of structure and stability from
    sequence is fundamentally a problem of physical
    chemistry.

8
Thermodynamics of membrane partitioning (1)
9
Thermodynamics of membrane partitioning (2)
  • Four step model
  • Partitioning, folding, insertion, and
    association.
  • Several pathways
  • Interfacial path, water path, combination of
    interfacial and water path.
  • Free energy of transfer or partitioning

10
Energetic components of partitioning (1)
Conformational Change.
Non-polar hydrophobic effect (expulsion of
non-polar compounds from water
Changes in motional degrees of freedom.
Difference in dielectric properties between water
and hydrocarbon region (mutual polarization
effects).
Direct electrostatic interaction between basic
residues and Anionic lipids.
Changes inside membrane.
11
Energetic components of partitioning (2)
  • Favorable partitioning of peptides and proteins
    into membranes is primarily caused by
  • Nonpolar interactions, e.g. hydrophobic effect.
  • Direct electrostatic interaction between charged
    amino acids and anionic lipids.

12
Energetic components of partitioning (3)
Solvation free energy
Non-classical hydrophobic effect, bilayer effects
Classical hydrophobic effect
for proteins
A is solvent accessible area
13
Stability of MP secondary structural elements (1)
  • Knowledge forces stabilizing soluble proteins
    comes from
  • Thermal unfolding experiments
  • Denaturation experiments
  • Computer experiments
  • MP display higher stability of their secondary
    elements in membranes
  • Strong resistance against denaturation and
    thermal unfolding.
  • The importance of Hydrogen bonding.

14
Stability of MP secondary structural elements (2)
Transfer free energy
DGt
Disruption of H-bond is unfavorable in
membrane e.g. 20 Residues would costs about 400
kJ mol-1.
non-H-bonded
Trp partitioning is favorable, but is not enough
to overcome non-H-bonded peptide bond DGt ? whole
residues can enter the HC core only if their
peptide bonds are H-bonded.
POPC interface more hydrophobic than water
Most hydrophobic side chain?
15
Molecular interpretation of DGt
  • Interpretation of DGt requires knowledge of
  • Structure of bilayer.
  • Transbilayer location of bound peptides.
  • Structure of peptides adopt.
  • Changes in bilayer as a result of partitioning.
  • Cellular membranes are in a fluid state
    (La-phase) for normal cell function.
  • High thermal disorder of membranes precludes
    high-resolution three-dimensional X-ray images.

16
Structure of fluid lipid bilayers
  • High thermal disorder reveals itself in
  • Width of atoms or atom groups distributions.
  • Combined thickness of interfacial regions is
    about the same as the HC core region
  • One interface can easily accommodate an
    (un)folded peptide.
  • Interfaces are extremely heterogeneous.

17
Distributions across bilayers
Distribution of atom groups experimental based
upon combination X-ray and neutron diffraction
methods (liquid crystallography).
Polarity, charge distribution peptide coming
from solvent undergoes dramatic variation in
environments polarity
(Un)folded peptide fits easily in interfacial
region.
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