Has anyone every heard of Schrondinger's equation

Frank Fuerst ffrank at rz.uni-potsdam.de
Wed Jan 31 11:55:22 EST 2001

Hi Dima,

klenchin at REMOVE_TO_REPLY.facstaff.wisc.edu (Dima Klenchin) wrote:

>Frank Fuerst <ffrank at rz.uni-potsdam.de> wrote:
>:It is generally asumed that proteins usually do indeed fold to the
>:global minimum of Gibb's free enthalpy, and this is experimentally
>:well supported for many of them (though there may be exceptions - but
>:I'm currently not aware of any).
>This is interesting. How can it be experimentally supported 
>without ability to calculate all possible minima and know exactly
>which one is global? 

Well, I didn't say "prove"...

The evidence is kind of indirect.
People have tried to start renaturation from many many different
conditions - and every time the same native fold was adopted (if it
was at all, I'll comment on your PIP kinase below). 
A different approach: In many proteins, mutations have been found to
have a great effect on folding kinetics and sometimes even on the
pathway, the intermediates and transition states involved etc. But
every time the protein folded to the same native structure as the wild

Surely proteins often get trapped in misfolded conformations, i.e.
conformations that are not the native fold. But it has been shown in
some cases (mainly by comparing them with equilibrium unfolding
intermediates), and is generally assumed, that these conformations
have a higher free energy at native conditions than the native fold.

If a protein is kinetically stable in a non-native form, either the
conditions are "not native enough", so the current conformation _is_
the lowest in energy under _these_ conditions, or this form is only
kinetically trapped.

I should point out again - as I did in the other posting,
<9597eq$fe2k0$1 at fu-berlin.de> answering Kresten, - that I don't argue
against the existence of different stable states under certain
circumstances, but against the concept that proteins fold under
kinetic control, to some energetic minimum that need not be the
global, but the fastest accessible minimum, governed by some specific
interactions in early intermediates.

For a review, see for example Jaenicke, R., Protein Folding: Local
Structures, Domains, Subunits, and Assemblies, Biochemistry 30 (1991),
3147-3158 (You see, the reference is rather dated - no one argues
about that now, as far as I see).

If there are strong interactions in the folding intermediates, this is
indeed often a problem on the way to the native fold, and this is why
so many, especially big proteins, have problems with refolding. 

>Sure, many small proteins do renature readily into native state,
>but still many do not. The bigger the size, the bigger the problem.
>Take actin. Not only it cannot be renatured, it cannot even be 
>folded properly in E.coli - there are _specialized_ actin-specific
>chaperones that fold it in the cell. 

I'm not familiar with the actin story, and chaperones also isn't my
special interest, but...

>I thought it is generally 
>accepted that chaperones are actively involved in the folding
>process of many (most?) eukariotic proteins. 

What does actively involved mean? Of course they are active:
They bind preferentially to partially folded proteins that still
expose a lot of hydrophobic surface, and they provide an environment
and/or energy to further _unfold_ these proteins. The concept is that
long-lived intermediates are often off-pathway intermediates, and the
protein needs to go up a bit on its conformational free energy
hypersurface to get to the right path again - the chaperones help them
doing this, they actively unfold them. I don't have a reference at
hand, but I would search for authors like Horwich and, ah look, I've
found one in my notebook:
Saibil H.
    Molecular chaperones: containers and surfaces for folding,
stabilising or unfolding proteins.
                       Curr Opin Struct Biol. 2000 Apr;10(2):251-8. 

>If so, I can 
>easily imagine that they burn ATP not only to accelerate reaction,
>but also to push it into alternative pathway. 

I've never heard or read someone reporting evidence for this in the
last years. 

The specificity of chaperones is because some of them are optimized
for binding of exposed parts of typical folding intermediates of their
target protein.

>Then there is my humble experience: PIP-5 kinase expressed 
>in E.coli in inclusion bodies "renatures" relatively easily into 
>some structure, which is perfectly soluble, compact (not a 
>random coil), but which has nothing to do with the native protein - 

This can be kind of molten globule, but

>no activity, no tri- and hexamers formation. 

If it ist "only" these properties that are changed, there might be
only very small structural differences to the native state, in a sense
that the perturbations are only locally, but nevertheless interfere
strongly with substrate and oligomer-partner binding.

>:>If so, then even if we solve all the math and computer problems
>:>that exist, we still cannot tell which one of many possible
>:>folds represents native protein. 
>:This shouldn't be a major problem.
>I think it is. As a rule, in the absence of crystal we have no clue 
>about structure based on sequence (homologs with structure
>don't count). 

This is because we can't calculate the lowest energy structure.

>Why do you think it isn't?

Because if we could calculate the lowest energy conformation, that
would be the native form, and because of evolution. Big proteins or
oligomers didn't fall from heaven, they evolved from smaller
precursors. And association or intramolecular domain-docking is
generally considerably slower than structure formation. Thus, for a
protein to evolve to an oligomeric or complex multidomain structure
consisting of parts that are *not* in their thermodynamic minimum,
imagine what evolution would have to do:
- Simultaneously mutate a protein in a way that it spends time in an
alternate conformation, 
- and mutate the other proteins (or the parts of the genome that are,
in the same step, moved, duplicated ... to form the other domains)
that they bind/dock to this conformation, forming a kinetically stable

>Yep, of course. The cross-talk is essential. My only point was
>that the bottleneck is at experimental stage now.

Nice to hear that, I'm an experimentalist in protein folding - but I
feel in the last decade progress has come from both sides: 
- Fast kinetics,
_ per-residue-resolution through mutagenesis or NMR 
and theory: 
- "theory in the computer", e.g. the Eaton et al. vs. Karplus et al.
story about beta-hairpin formation, see Dinner, Lazaridis, Karplus,
PNAS 96 (1999), 9068ff) 
- as well as theory "on the paper" (Dill, Wolynes ...)

Bye, Frank

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