Imperial College London

Henry Rzepa

Most scientific theories emerge slowly, over decades, but others emerge
fully formed virtually overnight as it were (think Einstein in 1905). A
third category is the supernova type, burning brightly for a short
while, but then vanishing (almost) without trace shortly thereafter. The
structure of DNA (of which I have blogged elsewhere) belongs to the
second class, whilst one of the brightest (and now entirely forgotten)
examples of the supernova type concerns the structure of proteins. In
1936, it must have seemed a sure bet that the first person to come up
with a successful theory of the origins of the (non-random) relatively
rigid structure of proteins would inevitably win a Nobel prize. Of
course this [did
happen](http://www.ch.imperial.ac.uk/rzepa/blog/?p=3621){target="_blank"}
for that other biologically important system, DNA, some 17 years later.
Compelling structures for larger molecules providing reliable atom-atom
distances based on crystallography were still in the future in 1936, and
so structural theories contained a fair element of speculation and
hopefully inspired guesswork (much as cosmological theories appear to
have nowadays!).

Dorothy Wrinch was a mathematician who came up with just such a
hypothesis for rigid protein structure, based in effect on elegance and
symmetry, coupled with some knowledge of chemistry and
crystallography\[cite\]10.1098/rspa.1937.0159\[/cite\]. She had noticed
that the repeating polypeptide motif might be folded such that a
cyclisation could occur to give what she termed a **cyclol** (an organic
chemist would call this an aminol, and we would also now recognize it as
a three-fold tetrahedral intermediate of the type involved in the
hydrolysis of peptides). Wrinch proposed that this cyclisation could be
repeated on a large scale to produce rigid scaffolds for proteins. The
three-fold symmetric elegance of such motifs clearly appealed to this
mathematician (the interesting symmetrical and conformational properties
of the central
[cyclohexane-like](http://www.ch.imperial.ac.uk/rzepa/blog/?p=1587){target="_blank"}
ring were still to be fully recognised by anyone. Since Wrinch built
many 3D models of her cyclols, one can but wonder how that central ring
was represented, and whether its chair conformation was at all
recognised. Another Nobel prize awaited the discoverer of this, Derek
Barton).

::: {#attachment_3747 .wp-caption .aligncenter style="width: 259px"}
![](http://www.ch.imperial.ac.uk/rzepa/blog/wp-content/uploads/2011/04/cyclol.jpg "cyclol"){.size-full
.wp-image-3747 .wp-caption .aligncenter loading="lazy" decoding="async"
aria-describedby="caption-attachment-3747" height="347" width="249"
srcset="https://www.ch.ic.ac.uk/rzepa/blog/wp-content/uploads/2011/04/cyclol.jpg 1246w, https://www.ch.ic.ac.uk/rzepa/blog/wp-content/uploads/2011/04/cyclol-215x300.jpg 215w, https://www.ch.ic.ac.uk/rzepa/blog/wp-content/uploads/2011/04/cyclol-734x1024.jpg 734w"
sizes="(max-width: 249px) 100vw, 249px"}

The Cyclol structure. Click for 3D.
:::

An immense controversy immediately broke out (not least because little
direct spectroscopic evidence for the OH groups could be found). The
story is rivetingly told by Patrick Coffey in his book *Cathedrals of
Science* (ISBN 978-0-19-532134-0). Linus Pauling entered the fray in
1939\[cite\]10.1021/ja01876a065\[/cite\], and one of the arguments he
deployed was not so much symmetric elegance but thermodynamics (he also
suggested hydrogen bonding and  S-S linkages for rigidifying proteins).
The proposed cyclisation, he suggested, led to a very high energy
species. Whilst Wrinch attempted to refute
this\[cite\]10.1021/ja01847a004\[/cite\], Pauling's arguments won almost
everyone over. Although Wrinch forlornly continued to promote her idea,
last reviewing the topic as late as in
1963\[cite\]10.1038/199564a0\[/cite\], crystallography was now producing
cast iron data for protein structures. None have ever emerged with a
**cyclol** motif, and this hypothesis is now firmly consigned to
untaught history\[cite\]10.1002/pro.5560060627\[/cite\]. To this day, no
examples of the *tris*(aminol) cyclol ring are to be found in the
Cambridge small molecule crystal structure database either, although
some related tetrahedral intermediates are known as crystalline species
(see for example
[here](https://wiki.ch.ic.ac.uk/wiki/index.php?title=Organic:arrow#Tetrahedral_intermediate_involved_in_Amide_hydrolysis.2FNucleophilic_addition){target="_blank"})
and they can be quite easily characterised in
solution.\[cite\]10.1021/ed064p725\[/cite\]

When I read the story, it struck me that modern theory could easily
verify how valid Pauling\'s thermodynamic argument was. I have picked
(ala)~6~ as my model, and have calculated the relative free energy
(ΔG~298~) of the following three isomers.

1.  An acyclic zwitterionic form of this hexapeptide, calculated with a
    SCRF reaction field for water to allow for the ionic nature
    (ωB97XD/6-31G(d,p)), reveals a proton transfer to a neutral system,
    with an energy of +7.3 kcal/mol  

    ::: {#attachment_3762 .wp-caption .aligncenter style="width: 234px"}
    ![](http://www.ch.imperial.ac.uk/rzepa/blog/wp-content/uploads/2011/04/model1.jpg "model1"){.size-full
    .wp-image-3762 loading="lazy" decoding="async"
    aria-describedby="caption-attachment-3762" height="174" width="224"
    srcset="https://www.ch.ic.ac.uk/rzepa/blog/wp-content/uploads/2011/04/model1.jpg 896w, https://www.ch.ic.ac.uk/rzepa/blog/wp-content/uploads/2011/04/model1-300x233.jpg 300w"
    sizes="(max-width: 224px) 100vw, 224px"}
    Acyclic (ala)6, in zwitterionic form
    :::
2.  A cyclic neutral peptide, which results from elimination of water
    from **1**, again calculated with a water reaction
    field\[cite\]10.14469/ch/8194\[/cite\], revealing a relative free
    energy of +0.0 kcal/mol  

    ::: {#attachment_3761 .wp-caption .aligncenter style="width: 232px"}
    ![](http://www.ch.imperial.ac.uk/rzepa/blog/wp-content/uploads/2011/04/model2.jpg "model2"){.size-full
    .wp-image-3761 loading="lazy" decoding="async"
    aria-describedby="caption-attachment-3761" height="178"
    onclick="jmolInitialize('../Jmol/');jmolSetAppletColor('white');jmolApplet([450,450],'load wp-content/uploads/2011/04/model2.cml;measure 6 48;measure 32 47;measure 36 47;measure 6 30;measure 3 33;zoom 100;spin 3;');"
    width="222"
    srcset="https://www.ch.ic.ac.uk/rzepa/blog/wp-content/uploads/2011/04/model2.jpg 888w, https://www.ch.ic.ac.uk/rzepa/blog/wp-content/uploads/2011/04/model2-300x239.jpg 300w"
    sizes="(max-width: 222px) 100vw, 222px"}
    Cyclic (ala)6
    :::
3.  The cyclic isomer **3** resulting from further cyclisation of
    **2**\[cite\]10.14469/ch/8197\[/cite\] with a relative free energy
    of +69.0 kcal/mol  

    ::: {#attachment_3763 .wp-caption .aligncenter style="width: 239px"}
    ![](http://www.ch.imperial.ac.uk/rzepa/blog/wp-content/uploads/2011/04/model3.jpg "model3"){.size-full
    .wp-image-3763 loading="lazy" decoding="async"
    aria-describedby="caption-attachment-3763" height="237"
    onclick="jmolInitialize('../Jmol/');jmolSetAppletColor('white');jmolApplet([450,450],'load wp-content/uploads/2011/04/model3.cml;measure 24 53;measure 16 19;zoom 100;spin 3;');"
    width="229"
    srcset="https://www.ch.ic.ac.uk/rzepa/blog/wp-content/uploads/2011/04/model3.jpg 914w, https://www.ch.ic.ac.uk/rzepa/blog/wp-content/uploads/2011/04/model3-289x300.jpg 289w"
    sizes="(max-width: 229px) 100vw, 229px"}
    Cyclol model for (ala)6.
    :::

From this, it appears that model **3** is \~69 kcal/mol less stable than
the cyclic peptide **2**, or 11.6 kcal/mol per amino acid residue.
Pauling\'s thermodynamic arguments suggested a value of \~28 kcal/mol
per residue (a value which Wrinch disputed as unreliable). So, in one
sense, the above calculation is closer to Wrinch than to Pauling! In
another, it still means Wrinch was wrong!! It is worth speculating why
Pauling\'s estimate is out. The cyclol **3** exhibits anomeric
stabilizations, which of course were unknown in Pauling\'s time. Both
**2** and **3** exhibit attractive, but different, van der Waals
attractions which contribute to their stabilities and Pauling took no
account of any entropy differences between **2** and **3.** In
retrospect,  **3** was [simply too
rigid](http://www.ch.imperial.ac.uk/rzepa/blog/?p=1174){target="_blank"}
to allow most enzyme catalysis models to function, as we recognise them
nowadays.

You might ask why I have revived a long forgotten theory as the topic of
this post. Well, I think it is always worth revisiting the past, and
re-examining old assumptions. When we do so, we find that Wrinch did not
miss by as much as her detractors perhaps implied. With a little more
luck, she might have gotten it right. Science is a bit like that, you
need a dose of luck sometimes!

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[https://orcid.org/0000-0002-8635-8390](https://orcid.org/0000-0002-8635-8390){target="_blank"
rel="author"}
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Most scientific theories emerge slowly, over decades, but others emerge fully formed virtually overnight as it were (think Einstein in 1905). A third category is the supernova type, burning brightly for a short while, but then vanishing (almost) without trace shortly thereafter.

Messaggi di Rogue Scholar

The Cyclol Hypothesis for protein structure: castles in the air.