Hydrophobic Mismatch between Proteins and Lipids in Membranes

Susanne Pfeifer tiffy@tiffy.it08.07.2004

Seminar Theoretical Analysis of Protein-Protein Interactions

Universität des SaarlandesChair of Prof. Dr. Volkhard Helms

2

Agenda

• Introduction• Possible adaptations to mismatch• Consequences of mismatch for:

• Proteins and peptides• Lipid structure and organization

• Effects of mismatch in biomembranes

3

4

Basics

Introduction

5

Basics

Introduction

6

Basics

Introduction

• Length of lipid-exposed hydrophobic segments is equal to the hydrophobicbilayer thickness

• Proteins that are encountered in one membrane can have different lengths of their hydrophobic parts

• Membrane proteins with the same length can be encountered in bilayers of different thickness

7

Basics

Questions

1. How do membranes deal with a mismatch between the hydrophobic part of a transmembrane protein and the bilayer thickness?

2. How important is the extent ofhydrophobic matching for membrane structure and function?

3. Could mismatch play a functional role?

8

Basics

Possible adaptations to mismatch • Positive mismatch

• The protein might oligomerize or aggregate in the membrane

to minimize the exposed hydrophobic area

• Transmembrane helices could tilt to reduce their effective hydrophobic length

• Transmembrane helices could adopt another conformation

9

Basics

Possible adaptations to mismatch• Negative mismatch

• Results in protein aggregation or changes in backbone conformation or side chain orientation

• Too short peptides might not incorporate and adopt asurface localization

• Lipids decrease the bilayer thickness by disordering their acyl chains

10

Basics

Implications for membranes

• Effects on • protein conformation• protein orientation• helical tilt• aggregational

behavior

can affect • protein activity• membrane insertion• protein assembly

• Effects on • lipid structure• lipid organization

have implications for • processes that are

sensitive to lipid packing

• Processes that require the local and transient formation of non-lamellar structures

11

Basics

Consequences of mismatch

• Consequences for properties of proteins• Protein activity and stability• Protein aggregation• Tilt• Localization at membrane surface• Protein/peptide backbone conformation

12

Basics - Consequences of mismatch

Protein activity & stability• The extent of hydrophobic matching is important

for determining the functional activity of proteins• There are a number of proteins that do not show

a clear optimum bilayer thickness for activity, but they require a minimal chain length

• many other factors may be involved in determining the functional activity of membrane proteins(e.g. lipid packing, fluidity, surface charge density, intrinsic curvature, lateral pressure profile, …)

Protein activity may be related to protein stability, which also can be affected by mismatch

13

Basics - Consequences of mismatch

Protein aggregation• Response to hydrophobic mismatch• Occurred only with a rather large mismatch:

• 4 Å thicker or• 10 Å thinner

than the estimated hydrophobic length of the proteinare allowed without induction of significant aggregation

• Proteins with long hydrophobic stretchtilt in the membrane Reduction of their effective length

• Comparison is difficult, because the lipids differ not only in acyl chain length, but also in other properties

14

Basics - Consequences of mismatch

Tilt

• Occurs if the hydrophobic part of a protein is too long to span the membrane

• Important for the functional and transport activity of membrane proteins

• An increase in helix tilt occurs at increasing protein content decrease in lipid order decrease in bilayer thickness

• Accompanied by a bend to reduce unfavorable effects on lipid packing

15

Basics - Consequences of mismatch

Tilt

Change in helix tilt change in protein activity

16

Basics - Consequences of mismatch

Tilt

• Special cases:• In large proteins:

changes in helical tilt have only little effect on lipid packing

• Single transmembrane helix: a tilt would cause a strain on the surrounding lipids to accommodate the helix in the bilayer

large degree of tilting is less favorable

17

Basics - Consequences of mismatch

Localization at membrane surface• Relatively small hydrophobic peptides

may not be able to integrate into the membrane• orientation at the membrane surface• Peptide aggregation outside the bilayer

• Amino acid composition is important (in determining the consequences of hydrophobic mismatch)

• The extent of membrane insertion for amphipathic pore-forming peptides is mismatch dependent

18

Basics - Consequences of mismatch

Localization at membrane surface• Surface-absorbed peptides insert their

hydrophobic side chains between the acyl chains near the membrane surface• membrane-thinning effect• dependent on the peptide/lipid ratio

• Important for studies • on the mismatch dependence of insertion for

such proteins• insertion of hydrophobic peptides with an

equilibrium between a transmembrane orientation and a surface localization

19

Basics - Consequences of mismatch

Backbone conformation

• Helix length fluctuates due to local variations in backbone structure

• Sensitivity of the backbone conformation for environmental changes depends on amino acid composition• Peptides with a hydrophobic stretch of

alternating leucine and alanine are more sensitive than peptides with a polyleucine sequence

20

Basics

Consequences of mismatch

• Consequences for lipid structure and organization• Lipid chain order• Phase transition temperature• Preferential interactions and

microdomain formation

21

Basics - Consequences of mismatch

Phase transition temperature• Melting transition temperature of lipid

bilayers is strongly affected• Proteins with long hydrophobic segments

stabilize the thicker gel phase• Short proteins stabilize the fluid phase

22

Basics - Consequences of mismatch

Microdomains

• In fluid bilayers consisting of lipids with different lengths, hydrophobic mismatch may induce preferential protein-lipid interactions formation of microdomains

• Systems consisting of two lipid species with different acyl chain lengths and one protein:hydrophobic mismatch induces preferential protein-lipid interactions(depending on hydrophobic length, differences in hydrophobic length…)

23

Basics

Effects in biomembranes

• Protein sorting• Membrane protein insertion and

topology• Regulation

24

Basics - Effects in biomembranes

Protein sorting

• Eukaryotic cell: • Level of cholesterol increases from the

endoplasmatic reticulum via the Golgi to the plasma membrane(suggesting a concomitant increase in membrane thickness)

• Protein sorting in Golgi is based on this length difference

• Increasing the hydrophobic length of proteins that normally reside in the Golgi

they can reroute the proteins to the plasma membrane (or vice versa)

25

Basics - Effects in biomembranes

Protein sorting

• Preferential protein-lipid interactions are consequences of hydrophobic mismatch results in domain formation and protein

sorting

26

Basics - Effects in biomembranes

Membrane protein insertion

• Signal sequences:• short hydrophobic length (7-15 amino acids)

• high tendency to form alpha-helical structures (with insufficient length to span a membrane)

• Length of signal sequences and mismatch are important for their functional activity

A mismatch could lead to a local destabilization in a bilayer

helps the translocation orpromotes preferential interactions with other short

helices of proteins in the translocation machinery

27

Basics - Effects in biomembranes

Membrane protein insertion

• Signal anchors• length closer to the hydrophobic thickness

of the membrane (19-27 amino acids)

influences the topology of proteins

• Stop transfer sequences hydrophobicity is more important than

length

28

Basics - Effects in biomembranes

Membrane thickness regulation

• A large variation in membrane thickness can be tolerated

• Variations of acyl chain length lead to changes in lipid composition • important for surface charge density• serves as tool to regulate local bilayer

thicknessprevention of unwanted consequences of hydrophobic mismatch in biological membranes

29

Basics

Results

• Hydrophobic mismatch • affects protein and lipid organisation• affects conformation and thermodynamic

properties of the membranes• plays a role in protein sorting in vivo• is required for specific

functional properties of membranes• depends on individual properties

30

Chain Packing

• Calculation of all possible lipid conformations• Probability of chain conformations

relative to their distances• Free interaction energy between

two inclusionsDetailed molecular-level information

on chain conformational properties

• Problems:• Computationally expansive• Full minimization of membrane

shape is difficult

31

Directors Model

• Theory-based model of elastic deformations is used to describe free energy differences associated with membrane perturbation due to protein-bilayer interactions

(Huang, 1986; Helfrich and Jacobsson, 1990; Nielsen et. al. 1998)

• All parameters were used beforein previous studies

• Thin, solvent-free lipid bilayer• With an embedded inclusion similar to a

gramicidin channel

32

Directors Model - Theory

The Model

33

Directors Model - Theory

The Model

34

Directors Model - Theory

Approximation of changes

• Elastic modes for approximation of changes in lipid packing:• Compression-Expansion (CE)

(due to changes in bilayer thickness)• Splay-Distortion (SD)

(due to variation in director among adjacent mol.)

• Surface-Tension (ST)(due to changes in bilayer surface area)

35

Directors Model - Theory

Total deformation free energy

• compression-expansion

rdrdr

du

dr

ud

dr

du

rKu

d

KG

r

r cdef

a

0

2222

02

14

rdrud

KG

r

rCE

a

0

2

02

4

rdrdr

ud

dr

du

rKG

r

r cSD

0

221

rdrdr

duG

r

rST

0

2

• surface tension

• splay-distortion

36

Directors Model - Results

Choice of boundary conditions• The bilayer deformation energy varies

as a function of• mechanical moduli• boundary conditions

• Problem:Energetic costs for packing the lipid molecules which are adjacent to the inclusion are not considered!

37

Directors Model - Results

Choice of boundary conditions

38

Directors Model - Results

Bilayer deformation profile• The shape of the deformation varies

as a function of the elastic moduli• Depending on the value of s, may the

bilayer deformation profile be nonmonotonic Energy minimization requirement may cause

a compression adjacent to the inclusion and an expansion further away from the bilayer/inclusion boundery

Packing Problemhydrophobic core volume per unit bilayer surface will deviate from its equilibrium value

39

Directors Model - Results

Bilayer deformation profile

rdrdr

du

dr

ud

dr

du

rKu

d

KG

r

r cdef

a

0

2222

02

14

40

Directors Model - Results

Bilayer deformation profile

rdrdr

du

dr

ud

dr

du

rKu

d

KG

r

r cdef

a

0

2222

02

14

41

Directors Model - Results

Bilayer deformation profile

rdrdr

du

dr

ud

dr

du

rKu

d

KG

r

r cdef

a

0

2222

02

14

42

Directors Model - Results

Radial decomposition of free energy• Depending on the choice of boundary

conditions GCE can be less, equal or largerthan GSD

• The relative contributions of these major components to Gdef vary in dependence of• s (contact slope) (length scale)

4

1204

1

a

c

K

dK

43

Directors Model - Results

Radial decomposition of free energy

44

Directors Model - Results

Radial decomposition of free energy

45

Directors Model - Discussion

Comparison

• The results of the presented model confirm and extend the findings of Huang (1986) and Helfrich and Jakobsson (1990)• Better results for s=0

• Failures with s=smin could arise because the parameters that are used may be inappropriateor additional contributions to Gdef which are neglected

• Today there is insufficient information to choose the appropriate boundary conditions

46

Directors Model - Discussion

Biological implications

47

Appendix

References

• Hydrophobic mismatch between proteins and lipids in membranes (1998, Killian)

• Energetics of Inclusion-Induced Bilayer Deformations (1998, Nielson et al)

• A Molecular Model for Lipid-Protein Interactions in Membranes: The Role of Hydrophobic Mismatch (1993, Deborah et al)

• Synthetic peptides as models for intrinsic membrane proteins (2003, Killian)

48