Membranes & Membane Proteins

8/13/2019 Membranes & Membane Proteins

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Membranes/membrane proteins

Importance of membranes and membraneproteins

General requirements for proteins to residein biological membranes

Different types of membrane proteins

-helical proteins Trans-membrane helix interactions


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Biological Membranes

Highly selective permeability barriers Amphiphilic organization: Hydrophobic in,

hydrophilic out Fluid mosaic model: liquid and asymmetric


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Importance of membranes

Compartimentalization: specific chemical environmentpH, redox potential,enzymatic composition, ioncomposition etc.


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Higher organization

Compartimentalization: specific chemical environmentpH, redox potential,enzymatic composition, ion composition etc.

Lyzosomes contains enzymes fordegradation of macromoleculesat low pH

Organization into organelles

Mitochondrion


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Membrane structures inorganelles

Inside mitochondria and chloroplasts further compartimentalization


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Importance of membranes

Chemical insulators: Charge gradient, energy generationProton gradient, energy generationIon gradients, signaling

Peter Mitchell's, Chemiosmotic Theory, Nobel prize 1978


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Chemiosmotic Theory


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Fundamental importance


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genomic composition: 20-30% of thegenes code for membrane proteins

pdb statistics, 25.000 structures250 membrane proteins


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Medical importance

Cytic fybrosis, chloride transporter Virus entry and maturation, receptors

Resistance against cytostatica, multi drugresistance proteins Bacterial infection, adhesins, transporters Certain cancer types Immune system Regulated cell death/apoptosis


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Economical importance

marketed small molecule drug targets grouped by class


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Fluid mosaic modelSinger and Nicholson, 1972


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Integral and PeripheralMembrane Proteins

Integral (yellow): bound by Hydrophobic interactions, can only beremoved by disruption of membranes, examples are cytochromeoxidase, GPCRs, channels.

Peripheral (blue): Bound by electrostatic and H-bond interactions,i.e. mitochondrial cytochrome c .


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Types of Membrane Proteins


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-helical and -barrelmembrane proteins

Photosystem I OmpF


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It costs energy to break hydrogenbonds in hydrophobic environments

It costs ~ 2 kcal/mol to bring one hydrogenbond from water into alkane. (C=O.H-N).

It costs ~6 kcal/mol to bring one hydrated pairof hydrogen bond donor/acceptor into alkane.(C=OH 2O; N-HH 2O).

It thus costs about 4 kcal/mol/peptide bond tomaintain unfolded/unsatisfied hydrogen

bonds in a membrane: It is impossible for anunfolded protein to reside in a membrane.


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Main-chain hydrogen bonds

membranehydrophobiccore

-helical proteins vs. - barrel proteins

The hydrogen-bonding patterns imply that -barrel proteins have to befolded prior to insertion into the membrane, whereas presence of2ndary structure is sufficient for helical proteins.


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Localization of and membrane proteins

-helical proteinsbacterial inner membranescell membranesER, golgi complexorganelle membranesinner and outer membranesof chloroplast andmitochondria

viruses

- barrel proteinouter membranes ofGram-negative bacteriaouter membranes ofchloroplasts and mitochondria


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Polytopic Membrane Proteins:Helical Proteins

Bacteriorhodopsin converts light into electrochemical energy. First structure of membrane protein, done by electron microscopy. 7 Transmembrane helices, almost perpendicular to membrane. Discussed in detail Bioenergetics


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Main Chain Hydrogen Bondsin -helices

Main chain hydrogen-bond donors andacceptors all participatein hydrogen bonds.

Helical membraneproteins are commonbecause it is easy tosatisfy hydrogen bond

requirements.

Side chains


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All side chains point outwardsinto membrane: They all need tobe hydrophobic.

helices are readily recognised insequence by stretches ofhydrophobic residues.

Side chainsin transmembrane -helices

Hydrophobic residues in yellow


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TM Amino acid composition

R K E D Q N H P Y W S T G A M C F L V I0

2

4

6

8

10

12

14

16

TM H. Sapiens (all)

Amino acid residue

a v e r a g e o c c u r e n c e

(adapted from Lui et al. Genome Biol 2002 and with data from Sanger institute)

Hydrophobic residues are overrepresentedCharged and polar residues are

under represented


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helical proteins Identification of helical membrane proteins Hydrophobicity plots

MALEPETVTVSEVVSPEYLDMRRRFWIALMLTIPVVILEMGGHGLKHFISGNGSSWIQLL

Kyte and Doolittle hydrophobicity scale-4.5 most polar, 4.5 most hydrophobicAla: 1.800 Arg: -4.500 Asn: -3.500 Asp: -3.500 Cys: 2.500 Gln: -3.500Glu: -3.500 Gly: -0.400 His: -3.200 Ile: 4.500 Leu: 3.800 Lys: -3.900Met: 1.900 Phe: 2.800 Pro: -1.600 Ser: -0.800 Thr: -0.700 Trp: -0.900Tyr: -1.300 Val: 4.200

- Take average hydrophobicity of 9 residues(a window) assign that to the central residue.- Shift the window by one residue etc.

Kyte, J. and Doolittle, R. 1982. J . Mol. Biol.


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Hydropathy plotsMALEPETVTVSEVVSPEYLDMRRRFWIALMLTIPVVILEMGGHGLKHFISGNGSSWIQLL


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TM regions

http://www.expasy.org/tools/protscale.html

7-8 predicted helices


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Number of TM regions

TM = Trans Membrane(Adapted from Krogh et al, JMB 2001)

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18012

3456789

1011121314151617

Num ber of helices

O c c u r e n c e

i n %

Predicted number of TMhelices in the E. coli genome


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Number of TM regions

(Adapted from Krogh et al, JMB 2001)

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 180

2.55

7.5

1012.5

1517.5

2022.5

2527.5

30

32.535

37.5

num ber of TM helices

o c c u r e n c e

i n %

Predicted number of TM helicesin the E. coli , C. elegans genome


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Interface residues

Aromatic residues (Trp, Tyr

and Phe) are abundant at thepolar/apolar border

Tyrosine hydroxyl pointoutward


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Helix angles

bacteriorhodopsin


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Helix angles

Van den Burg et al. Nature 2004

SecYEG complex

~0-35 o tilt

avarage~ 21 o


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The hydrophobic effect

The hydrophobic effect gives soluble proteins their compactness.

aqueous

solvent

hydrophobic

polar

ordered solvent

b h l


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Interaction between helices

What then gives TM proteins their compactness and stability?

aqueous

solvent

hydrophobic

polar



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Polar interactionsFew polar residues in TM regions

Those that are often involved in inter-TM hydrogen bonds

Rhodopsin

Ubarretxena-Belandia and Engelman, Curr. Op. Struc. Biol. 2001



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Glycophorin ATM most often form coiled-coils (left-handed, right-handed)

Knob into holes packingby hydrophobic residues

Close VdW interaction packing

Protein-protein packing moreefficient that lipid protein packing

LIxxGVxxGVxxT

MacKenzie et al. (1997) Science 276 , 131-133.

Helical wheel


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motifHelix 3.6 residue per turn

Residue n and n+4 point inabout same direction

Heptad repeat, 7 residues



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GXXXG motif

LIxxGxxxG VxxT in glycophorin A

GXXXG (or GG4) very close packing

Formation of C -H --- O hydrogen bonds

In hydrophobic environment C -H --- Oabout half strength of ordinary NH --- O

MacKenzie et al. (1997) Science 276 , 131-133.


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Identif ing interactions: TOXCAT


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Identifying interactions: TOXCAT

TOXCAT system: MBP domain-membrane helix-ToxR domain

Oligomerization activates ToxR

Oligomeric ToxR controls expressionof chloramphenicol-acetyl transferase(CAT)

CAT gives resistance againstchloramphenicol Giving resistanceagainst

Russ and Engelman PNAS 1999


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Other Motifs


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Other Motifs

SxxSSxxT

SxxxSSxxT

GxxxxxxG

AxxxA

Monotopic membrane proteins


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Monotopic membrane proteins

Are embedded in but do not traverse themembrane completely

Example: Prostaglandin N-synthase


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F i P l di S h


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Function Prostaglandin Synthase

Catalyses synthesis ofProstaglandin H 2 fromarachidonic acid.

The fatty acid is ahydrophobic degradationproduct of lipids.

Prostaglandins are signal

molecules


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Summarized


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SummarizedImportance of membrane proteins

Hydrogen bonds satisfied within hydrophobic part of membranes

Side-chains of TM a-helix hydrophobic

Interactions between TM-helices- Polar interaction side chains

- VdW interactions due to close packing

- C-H...O hydrogen bond

Monotopic membrane protein architecture

Membranes & Membane Proteins

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