Lecture 8: Predicting/Testing Membrane Protein Structure/Topology Flashcards
What are amino acids joined by?
Polar Peptide bonds formed in a condensation reaction
What is the exceptional AA?
Proline is an imino acid
What are the amino acids in the alpha helical parts?
Non-polar/hydrophobic
What is not possible in the TM regions?
Hydrogen bonding to water as there is none
What neutralises the charge?
H bonds between the peptide bonds
What is the pattern?
Regular H bonds between CO delta minus and NH delta plus. Between 1-4, 2-5, 3-6 etc forms alpha helix
What do the R groups do?
Project out from the helix and are not involved in H bonding
How many residues are there per turn at what length, what length per residue?
3.6, 5.4 A and 1.5 A
How many AA are needed to span the bilayer in helical conformation?
20
What is the length of the hydrophobic core of the bilayer?
30A
Describe the basic structure of B-sheet membrane proteins
Every other AA R group projects in opposite orientation. Regular H bonding patterns between different strands. Each peptide bond is H bonded to another peptide bond in a different sheet. All the hydrophobic AAs could face outwards and all the hydrophilic ones could face inwards.
How is an antiparallel B sheet formed?
with adjacent B strands running in opposite directions
What is the smallest B barrel?
OmpX
How many AA are needed to span the membrane with a B barrel and why is this different to alpha helix?
8-9 because in a more extended conformation
What is alternated?
Whether they are lipid exposed or internal
What are the inside facing lipids?
Hydrophilic and polar
What does OmpX act as and how?
An adhesion protein as the protruding loops provide binding sites for other proteins with B strands
What are the different hydrophobicity scales?
Kyte, Doolittle and Engelmann and Steitz
How can you predict whether a membrane protein is likely to be integral?
By assigning each AA a hydrophobicity value and averaging over 15-20 AA.
What does the hydrophobicity analysis allow?
Prediction of the number of TM spanning segments
What is plotted?
The averages after moving the window along by one AA at a time
Why can this method be difficult?
The cut off for a hydrophobicity TM domain can be arbitrary eg -1
What does hydrophobicity analysis not work with and why?
B barrels as the hydrophobic/hydrophilic regions are alternate so are cancelled out
Why is a smaller window not used?
Gets a very messy plot as the averages are more greatly affected by shifting the window by 1.
Why aren’t all TM helices hydrophobic and give an example?
Not all hydrophobic stretches are part of the TM domains eg. Lac Y where some hydrophilic domains are part of the helix for protein-lipid interactions rather than protein-protein interactions.
What two ways can the predictions be improved?
The positive inside rule and sequence alignments
What is the positive inside rule?
Cytoplasmic loops of membrane proteins contain lots of positively charged AAs compared to extracytoplasmic loops. The positive regions are usually positioned a certain distance fro, the TM region so longer loops may not be positive far away
What does the TOPPRED programme do?
Takes into account hydrophobicity and the positive inside rule to predict the number and orientation/topology of the protein.
What can be positionally conserved?
AAs
Which AAs are frequent in TM regions?
Leucine and isoleucine
What different contacts can alpha helices make?
Protein-protein or protein-lipid
What AA sequences are common for knobs into hole interaction?
GXXXG, GXXXA promotes interactions between helices
What is the pattern of these sequences?
The GG and GA parts will be on the same face of the helix.
In bovine cytochrome C oxidase, what is positioned along the helix so they can interact with each other via H bonds? What is the pattern?
Serines. They are positioned 2 turns of the helix from each other.
Conserved H-bonds between polar residues in TM segments are important for what?
helix alignments
