| Chem 431 |
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Fall 2001 |
| Lecture Notes:: 24 September |
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| PREVIOUS |
Let's look at folding again: Would guess protein would fold to lowest free energy conformation. Problem: is there time? ("Levinthal's Paradox", formulated by Cyrus Levinthal in 1968) Stryer calculation (very conservative): Assume 100 aa residue protein with 3 possible conformations/residue; then get 3100or 5 x 1047 possible conformations. If search at a rate of one structure/10-13sec then get (5 x 1047)(10-13)= 5 x 1034 sec or 1.6 x 1027 years to search (and thus to fold protein). This is greater than the age of our Universe (10-15 x 109 yrs). Rawn calculation (perhaps more realistic): same but assume 10 conformations, then get 1087sec or 3 x 1080 yrs!
Obviously from these calculations not searching all possible conformations (or we have the process wrong!), so cannot say protein achieves the lowest global free energy, but rather a local free energy minima. (Like a valley in mountain range: a local energy minima, but not lowest [Marianas trench].) Which valley the protein reaches will depend on folding "paths." That is folding appears to depend on kinetics as well as thermodynamics.
Some structures fold faster than others: alpha-helix seems fastest folding, due to cooperativity. That is, once a couple of H-bonds of helix are formed subsequent residues are aligned to form H-bonds, thus a cooperative process. Beta-structures are second most rapidly formed. Thus commonness of these two structure types.
From our discussion last time we expect rapid formation of alpha and beta structures. However, unless they are stabilized by interacting with each other, they will unfold and try other combinations until stable associations result. (G&G, Figure 6.36, p 193) [overhead, V&V 8.5].
Note that some aa residues favor one or another secondary structure (G&G, Table 6.3, p 198). Unfortunately, such tendencies by themselves have not proved effective in predicting protein structures. However, using this kind of information in a "local" (nucleation) and "hierachical" (extensions/higher levels) way can predict some small protein structures reasonably well, as in the LINUS computer program discussed in G&G, pp 198-9.
Now known that many proteins are aided in folding process by Chaperones: appear to stabilize unfolded conformation, allowing time to find correct folding pattern. Some chaperons are known to have a barrel-shape into which new or partially denatured protein is inserted, native protein is released. Chaperones require ATP energy to function. The so-called Heat-shock proteins are a family of chaperons.
"Breathing" motions:
Myoglobin: Myoglobin is a globular protein in the globin family. Eight alpha helices in tertiary structure; about 80% alpha helix (high for globular proteins). (G&G, Figure 15.24, p481) [overhead 7-41, V&V] Interior almost exclusively hydrophobic residues, with water excluded from interior. Surface has mix of hydrophobic and hydrophilic residues, with ionizable groups on surface. Note myoglobin is doing for oxygen binding.
Myoglobin functions to store and facilitate the diffusion of oxygen in muscle. Oxygen binds to a heme {Fe (II)-protoporphyrin IX} prosthetic grp. Four of irons six ligands are to heme nitrogens, with a fifth to his nitrogen. The final ligand bond goes to oxygen. (G&G, Figure 15.25, p 482) [overhead, S, Figure 7-5, 7-8] Breathing motions are necessary to allow the exchange of oxygen, since the heme is in a closed pocket. [overhead 8-9, 8-10, V&V]
Hemoglobin: Hemoglobin is an aabb oligomeric protein: its quaternary structure consists of a tetramer of myoglobin like subunits. (G&G, Figure 15.24, p481) [overhead 9-13a, V&V] The two types of chain are slightly shorter than myoglobin chains (a= 141 aa residues, b= 146 aa residues). There are extensive contacts between an a and a b subunit to give a dimer. The dimers have additional contacts to give the tetramer. Oxygen binding results in a change of conformation in Hb. (G&G, Figure 15.31, p485) [overheads 9-13a vs. 9-13b, V&V] The change of conformation affects the binding of oxygen (G&G, Figure 15.32, p486) [overhead Fig 9-16, V&V] {oxygen binding is reduced in the "blue" form due to steric hindrance between the oxygen and the heme}.
Let's look at binding in terms of saturation, Y, where if Y = 1 every site of every Myoglobin is occupied by an oxygen molecule (thus if Y = 0.5, then 50% of the myoglobin are binding oxygen and 50% are "empty"). Mb/Hb binding curve [overhead 33 V&V]:
Can describe binding as dissociation equilibrium, then: 
& 
for saturation. Substituting, 
,
the equation of a hyperbola. If expressed as pressures, then 
where P50 = p2 @ 50% saturation.
Note that the binding curve for Mb is indeed hyperbolic in shape.
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