Protein Folding
process by which a polypeptide chain acquires its 3dimentional biologically active structure (native state)
Can involve enzymes which catalyse the formation and exchange of disulphide bonds or involve chaperones which bind partly folded polypeptide and prevents it making illicit associations with other proteins
Chaperones also promote folding
Molten globular state - looser tertiary structure than the native state after most secondary structures have formed successfully
Major unsolved problem in protein structural biology - how primary sequence relates to tertiary structure
Proteins are not static in their native state
Two major contributors
Enthalpy
Entropy
All non covalent interactions
energy is required to create order.
difference between enthalpy and entropy is the free energy of the stable state
More known about factors influencing stability of the native state of a protein compared to the unfolded state
the marginal difference between native and unfolded states is biologically very important especially for globular proteins which have a high turn over rate. i.e it has to be just as easy to degrade a protein as to synthesise one. Furthermore the catalytic activity of enzymes requires a large degree of flexibility to accommodate conformational changes and binding to ligands which would otherwise be inconceivable in a rigidly stable structure
Only one conformation with significantly lower free energy than all other possible conformations
How is folding achieved on such a short timescale
Difficult to examine experimentally due to the transient nature of the intermediates
If kinetic factors are important for folding it is possible the final folded state is not the most energetically favourable but, just the most stable of the conformations that are accessible
Example by replacing of six residue loop region (important for function) with shorter ones with a propensity to form beta turns - protein folded quicker, indicating that by selecting for function evolution has ultimately compromised the speed of folding and the stability of the native state
Most common obsticles (1) aggregation of intermediates (2) formation of incorrect disulphide bonds (3) isomerization of prolines
Other ways to remove kinetic barriers include the removal of segments of proteins that help the remaining structure to fold and form the active form of the protein
Chaperones
Proteins are metastable
Little change in hydrogen bonds forming secondary structures because they form just as easily with water molecules which are equally stable - so cannot be driving force of folding
Instead large changes in free energy occur when hydrophobic residues out of water and into contact with one another in a hydrophobic core
Significantly reduces number of possible conformations because only those sterically accessible are sampled
Some hydrophobic sidechains are buried whilst others are exposed on surface
Driving force for protein folding
NH and CO groups buried in hydrophobic core is unfavourable so formation of secondary structures early in the folding process can be attributed to burying of hydrophobic residues
single and multiple pathways observed e.g. barnase and lysozyme respectively
Techniques to measure protein folding
Enzymes can help with disulphide bond formation during folding
If the second residue is a proline then the cis conformation of the peptide bond is only about 4 times less stable than the trans increases it's occurrence in proteins
Isomerization of proline residues can be a rate limiting step especially in vitro
In vivo catalysed by enzymes called peptidyl prolyl isomerases
Involved in immunosupression by inhibiting prolifiration of T cells
Bind immunosuppressive drugs
Mechanism?????
Protein folding and Disease
cassette tape analogy