Membrane Proteins 1/2 Flashcards
How can membrane proteins act as a means by which information can enter a cell?
Change conformation upon ligand binding and transmit a signal across the membrane.
How are high levels of complexity achieved in membrane proteins?
Via association with different proteins and other molecules.
Give some molecules that associate with photosystem II?
Carotenoids, quinones, chlorophyll etc.
Describe the composition of a membrane.
Made of a phospholipid bilayer, complex mix of lipids and membrane proteins.
How are lipids within the membrane affected by hydrophobicity?
Hydrophobicity often dictates lipid position and function.
What regions of a membrane protein are associated with function?
Soluble regions
What are multipass proteins?
Made of several integral membrane proteins that pack together to allow the overall function of the protein.
How can a peripheral membrane protein be anchored to a membrane?
By an amphipathic helix or a lipid
What is an amphipathic helix?
One side of the helix is hydrophobic and the other is hydrophilic, allowing insertion into the membrane.
Which lipid modifications of membrane proteins occur on the intracellular side of the membrane?
Palmitoyl, N-myristoyl and Farnesyl additions.
Where are GPI anchors found and what do they consist of?
Only added on the extracellular side of the membrane. GPI anchors consist of phosphate, sugar and lipid groups.
What is a monotopic membrane protein?
A small region of the protein inserts into the membrane but does not pass all the way through.
Give an example of a monotopic membrane protein.
Electron transfer flavoprotein-ubiquinone oxidoreductase which must associate with the membrane in order to access ubiquinone which is lipid soluble.
Describe a beta barrel membrane protein.
Beta strands making up a barrel where hydrophobic residues face the lipid environment and hydrophilic residues face the hydrophilic core and interact with charged substrates. Allows formation of an aqueous pore through the membrane.
Give examples of beta barrel membrane proteins.
FepA/OmpLA- iron transport
Maltoporin- maltose transport
TolC- drug export from bacteria
Describe the features of an alpha helix.
3.6 residues per turn, stabilised by intrahelical hydrogen bonding between n and n+4 residues. All hydrogen bond donors and acceptors are used- very stable.
How else can alpha helices be stabilised in a membrane protein?
Stabilised by hydrophobic interactions between side chains that project out from the helix.
What is helix length partially dependent on?
The lipid composition of the membrane- areas including lipid rafts will be thicker and require a longer helix to cross the membrane.
What is given in a hydrophobicity scale?
The free energy required to transfer each amino acid from a lipid to an aqueous environment. If less energy is required, the amino acid is more likely to exist in an aqueous environment.
Which amino acids are more likely to exist in an aqueous environment?
Charged amino acids that can form polar/hydrophobic interactions with other molecules in the aqueous environment.
Which amino acids are more likely to exist in the membrane environment?
Amino acids which require a lot of energy to be transferred to an aqueous environment, e.g. Phe/Met
Why do TM domains carry some hydrophilic and charged residues? Give an example.
To allow the formation of pores, necessary for transport. An example is the K+ channel formed from helices creating a helix bundle.
What is hydrophobicity analysis?
Prediction of the number and location of TM domains from the primary sequence of a protein.
Why is topology important for membrane proteins?
Membrane proteins are asymmetrical and must be guided into the membrane in the correct orientation in order to function properly.
Give some functions of membrane proteins.
Produce oxygen, use oxygen to produce energy, ion and nutrient transport, removal of waste products/toxins from cells.
How was it originally suggested that proteins could exist in both orientations?
Originally thought that there were two genes coding for the protein with different topology and charge distribution.
What was proven to be the way proteins can exist in both orientations?
One gene coding for one protein, with little charge difference between inside and outside regions so can exist in either orientation.
What is meant by the positive inside rule?
Membrane proteins tend to have more positively charged residues in regions on the intracellular side of the membrane- this maintains asymmetry.
Give programs that can predict the orientation of alpha helices.
TMHMM and Prodiv-TMHMM
How does TMHMM predict the orientation of alpha helices?
TMHMM searches for continuous stretches of hydrophobic residues and looks at the distribution of positively charged residues in flanking regions.
What are the limitations of TMHMM?
Cannot predict localisation of soluble proteins or the topology of beta barrel proteins.
Why can’t TMHMM predict the topology of beta barrels?
A beta barrel has a completely different distribution of charged amino acids to an alpha helix.
How can topology be determined experimentally?
Generation of fusion proteins, using a marker that only folds or gives a signal in a specific part of the cell. Generate fusion proteins with the markers in different places in order to determine the location of the termini and loop regions- allows determination of the orientation of the protein in the membrane.
How can GFP and PhoA be used to experimentally determine topology?
GFP- only folds and fluoresces in the cytosol
PhoA- only folds in the periplasm between the membrane
Regions that show PhoA activity shouldn’t give GFP signal.
Describe a 3(10) helix?
Hydrogen bonds between n and n+3 residues. Forms a longer and thinner helix. Less stable than standard alpha helix due to hydrogen bond donors/acceptors not all being used.
Describe a pi helix.
Hydrogen bonds between n and n+5 residues. Forms a shorter and fatter helix. Less stable than standard alpha helix due to hydrogen bond donors/acceptors not all being used. Can have a region of pi helix within a standard alpha helix- called a pi bulge.
What do helix variations cause?
Regions of local instability which is often important for conformational change. Allows for similarly structured proteins with different functions.
What is the role of proline residues within a membrane protein?
Acts as a helix breaker, forces polypeptide chain into a conformation that isn’t ideal for helix formation, causing a random coil region to form.
What is the role of glycine residues within a membrane protein?
Allows tight packing of the helix and provides a flat surface for helix packing. When found either side of a TM domain, it allows the next TM domain to pack closely to the first. Can also induce local flexibility.
What is the aromatic belt in membrane proteins?
Collection of Trp/Tyr residues at the transition point between the non-membrane and membrane environment as these residues can exist in both environments.
Which lipid is essential for the folding of lactose permease?
Phosphatidylehthanolamine (PE)
What is the role of cholesterol in the membrane?
Reduces permeability and increases the rigidity of the membrane. Involved in post-Golgi protein sorting. Necessary for lipid raft assembly.
How can cholesterol affect membrane proteins?
Can stabilise certain conformations. Increases thermostability of MPs. Can affect ligand binding affinity of MPs.
How does cholesterol bind to membrane proteins?
Via a groove on the surface of the protein.
For what family of proteins is cholesterol essential for function?
GPCRs
Why is cholesterol essential in macrophage membranes?
Allows macrophages to squeeze through layers of epithelial cells without bursting.
