1. The second large class of proteins distinct from globular proteins are the membrane proteins.
It is estimated that 20-30% of all genes in most genomes encode for membrane proteins.
They are also the target of over 50% of all modern medicinal drugs.
Introduction
The first membrane protein to be
sequenced was Glycophorin.
2. The first membrane protein structure to be solved was of the bacterial photosynthetic reaction
center from Rhodopseudomonas viridis by Hartmut Michel, Johann Deisenhofer and Robert
Huber for which they shared the Nobel Prize in Chemistry for the year 1988.
3. Membrane structure and proteins
The exterior membranes
contain many pumps,
channels, receptors and
enzymes and the protein
content of is typically 50%.
4. Energy-transduction membranes such as the
internal membranes of mitochondria and
chloroplasts have the highest content of protein,
typically 75%.
5. Genomes and membrane proteins
Wallin & Heijne, 1998, Protein Science , 7, 1029-1038.
6. The survey of the Human Transmembrane Proteome (UCSF)
33,610 protein sequences in the human genome
29,375 unique protein sequences (no alternative splicing).
7,299 protein sequences are predicted to have at least one transmembrane helix (25%).
3,838 protein sequences predicted to have at least two transmembrane helices (13%).
3,418 unique sequences after removing all residues before and after the first and last
predicted TMH residue.
2,926 unique sequences after clustering at 98% sequence identity.
8. 3D Structure
As of January 2013 less than 0.1% of protein structures determined were membrane proteins
despite being 20-30% of the total proteome.
9. RCSB statistics of Membrane structures
Membrane 6727
Membrane part 4378
Membrane-enclosed lumen 1562
Channels/Pores 858
Electrochemical Potential-driven transporters 159
Primary active transporters 735
Group translocators 41
Transmembrane electron carriers 39
Accessory factors involved in transport 224
incompletely characterized transport system 43
10. Function
Membrane proteins perform a variety of functions vital to the survival of organisms:
Transport
Signal transductionEnzymatic activity
Cell-cell recognitionIntercellular joining Attachment to the cytoskeleton
and extra--cellular matrix (ECM)
11. It is possible to identify membrane proteins by the distribution of residues with hydrophobic
side chains throughout the primary sequence.
Membrane protein prediction
Early prediction of TM segments for helical IMPs generally used the following four-step
procedure
1. Derive propensity scale, a set of 20 numbers corresponding to properties or statistics of the
20 amino acids when found in TM regions.
2. Generate a plot of propensity values along the query sequence.
3. Smooth the plot by taking the average propensity value in a window of N residues and
plot the average at the center of the window (i.e. a sliding window average).
4. Identify TM stretches on the smoothed plot using some propensity threshold.
12. Amino acid Parameter Amino acid Parameter Amino acid Parameter
Ala 1.80 Gly -0.40 Pro -1.60
Arg -4.50 His -3.20 Ser -0.80
Asn -3.50 Ile 4.50 Thr -0.70
Asp -3.50 Leu 3.80 Tyr -0.90
Cys 2.50 Lys -3.90 Trp -1.30
Gln -3.50 Met 1.90 Val 4.20
Glu -3.50 Phe 2.80
Kyte and Doolittle scheme of ranking hydrophobicity of side chains
13. Hydropathy plot
Hydropathy plots reflect the preference of amino side chains for polar and non-polar
environments. The hydropathy values reflect measurements of the free energy of transfer of an
amino acid from non-polar to polar solvents.
Modern approaches based on Hidden Markov Models (HMMs) or Neural networks (NNs) are
related to sliding-window hydropathy plot methods.
15. Integral membrane proteins
They can be classified according to their relationship with the bilayer:
IMPs are permanently attached to the membrane and can be separated from the biological
membranes only using detergents, nonpolar solvents or sometimes denaturing agents.
16. 2. Integral polytopic IMPs span across the
membrane at least once. They have one of
two tertiary structures, α bundle and β
barrel (which are found only in outer
membranes of Gram-negative bacteria,
lipid-rich cell walls of a few Gram-positive
bacteria and outer membranes of
mitochondria and chloroplasts).
1. Integral monotopic IMPs are attached to
only one side of the membrane and do not
span the whole way across.
17. PMPs are temporarily attached either to the lipid bilayer or to integral proteins by a
combination of hydrophobic, electrostatic and/or other non-covalent interactions. Peripheral
proteins dissociate following treatment with a polar reagent, such as a solution with an
elevated pH or high salt concentrations.
Peripheral membrane proteins
PH domain of phospholipase C δ 1
18. These are located outside the lipid bilayer, on either the extracellular or cytoplasmic surface,
but are covalently linked to a lipid molecule that is situated within the bilayer.
Lipid-anchored proteins
19. Folding of Membrane Proteins
bovine rhodopsin human mitochondrial voltage-dependent
anion channel
α-helical bundles are structurally and functionally more versatile, serving as receptors,
channels, transporters, electron transporters, and redox facilitators.
β-barrels, made up of β-strands, are found in Gram-negative bacterial outer membranes as
well as in mitochondrial and chloroplast membranes, and these structures function as channels
or transporters for nutrients, proteins, hydrophobic toxic substances, and other molecules.
20.
21. How do their general structural features compare with those of soluble proteins?
It is almost same. The interior amino acids are found to be almost exclusively nonpolar and
packed just as tightly as those of soluble proteins, as suggested by measurements of the partial
specific volume of bacteriorhodopsin.
Because of the length and the highly nonpolar character of TM helices, hydropathy plots have
proven to be extraordinarily useful and remarkably accurate for predicting the topology of α-
helical MPs.
However, the amino acids of these outer surfaces are more hydrophobic.
The average lengths of the traversing secondary structure elements are greater than for soluble
proteins so that the 30 Å thick bilayer core can be spanned (α-helices are generally longer than
20 amino acids and β-strands longer than 10 amino acids).
22. Estimating the Molecular Weight of Membrane Proteins
SDS – PAGE is generally used to measure the molecular weight of soluble proteins.
In contrast to the relatively unstructured SDS-induced unfolded states of most water-soluble
proteins, unfolded states of membrane proteins contain a significant percentage of secondary
structure.
It has been observed that native (e.g., glycophorin A) or irreversible (e.g., GPCRs)
oligomerization may occur in SDS micelles.
Also, if not boiled prior to electrophoresis, OmpA migrates to different positions on SDS–
PAGE depending on the compactness of its structure (e.g. native OmpA migrates with an
apparent molecular weight of around 30 kDa, whereas completely unfolded OmpA migrates
as an around 35-kDa protein).
Thus, in some cases, differential binding of SDS to membrane proteins during electrophoresis
has been suggested to cause deviations up to 50% in their apparent molecular weights.
23. Amino Acid Composition
Leu, Ile and Phe residues are the mostly abundant in the acyl chain areas of membrane lipids.
Lys and Arg residues, with their long and flexible side-chains, are found at the lipid/water
interface region facing cytoplasm (Positive-inside rule). They neutralize their positive charge
by interacting with the negatively charged phosphate groups of phospholipid.
Trp and Tyr residues in the interface and polar residues in the aqueous zone. Trp and Tyr
residues have such a marked tendency to locate in the interfacial area that most membrane
proteins have what is known as an “ aromatic belt. ”
24. Membrane proteins in Drug Discovery
The G protein-coupled receptors (GPCRs) are the largest, most versatile, group of membrane
receptors and also the most pharmaceutically important, accounting for over 50% of all
human drug targets and acting as therapeutic targets for a wide range of disease conditions
including cancer, cardiovascular, metabolic, CNS and inflammatory diseases.
Ion channels represent another group of important membrane protein drug targets and account
for the activity of 10% of the currently marketed drugs.
Drews, 2000, Science, 287, 1960-1964.
25.
26. The European Membrane Protein Consortium (http://wwww.e-mep.org)
Japan Biological Information Research Center, Tokyo (http://www.jbic.or.jp/bio/english/)
Centers for Innovation in Membrane Protein Production, UCSF, Scripps ()
Membrane proteins of known structures
(http://blanco.biomol.uci.edu/Membrane_Proteins_xtal.html)
Transport DB (http://www.membranetransport.org)
