SFP16: Membrane Proteins

Question 1

What do we mean by ‘resolution’?

Answer 1

Resolution in protein structure is a measure of the detail of the data

Question 2

What would a high resolution (1-2A) mean?

Answer 2

Highest level of detail, will know positions of individual atoms

Question 3

What would a medium resolution (3-5A) mean?

Answer 3

Would know roughly where an amino acid side chain but not with enough precision to say this side chain must be interacting with this side chain etc.

Question 4

What would a low resolution (>5A) mean?

Answer 4

Would show very little detail at the amino acid level?

Question 5

Why is the protein data bank skewed?

Answer 5

• 25% of all genes encode membrane-spanning proteins

But <1% of proteins in the PDB

Question 6

why is NMR spectroscopy limited?

Answer 6

IT is limited to something very small (<30kDa) but most proteins are larger than this

Question 7

What is the example of bacteriorhodopsin converting radioactive energy into chemical energy?

Answer 7

• Under high light intensity huge amount of bR are synthesised by the light-harvesting bacteria, Halobacterium halobium
• light promotes a trans-> cis isomerisation of a bound retinal in bR
• Protons are pumped from the cytosol to the extracellular space creating a proton gradient
• proton gradient drives ATP synthesis by F-type ATPases

Question 8

How does electron microscopy with 2D crystals give 3D data?

Answer 8

EM is a low signal:noise technique i.e. each individual protein molecule can be ‘seen’ but the image is in distinct, low resolution
However all particles in a 2D crystal have the same orientation, you can ‘average’ the data from millions of protein particles -> higher resolution

Question 9

What is the electron microscope - better resolution?

Answer 9

Enables structural determination at almost atomic resolution (certainly 2.5A)

• tolerant of sample impurities
• does not require crystallisation (this is a big deal, as growing protein crystals is not a precise science)