Pasteurized Crystals – Tartaric acid.

What is it?

December 27 marks the 192nd birthday of Louis Pasteur, which means that (a) he’d be really old if he hadn’t died in 1895, and (b) today is the perfect day to talk about tartaric acid.

Tartaric acid occurs naturally in many plants, particularly grapes. You’ve already read about ‘wine diamonds’ (potassium bitartrate), but you may not be aware of the contribution tartaric acid has made to scientific language.

Naturally occurring tartaric acid, first isolated in 1769, was found to rotate plane polarized light to the right. When it was prepared synthetically, tartaric acid had identical properties, except that it didn’t rotate plane polarized light. The synthetic material was thought to be a different compound, and was named racemic acid (from racemus, Latin for ‘a bunch of grapes’). It was subsequently determined that tartaric acid can exist in two different forms; and that naturally occurring tartaric acid was L-tartaric acid, while ‘racemic acid’ was actually an equal mixture of D and L-tartaric acid, mirror image isomers (enantiomers). These enantiomers were optically active in opposing directions, appearing optically inactive; this explained the otherwise identical properties of tartaric acid and racemic acid.

For this reason, ‘racemic’ came to mean ‘an equal mixture of enantiomers’, and this term continues to be ubiquitous in organic chemistry today.

So where does Pasteur fit into this story? Early in his career, before he discovered vaccination, microbial fermentation and invented the process which still bears his name (pasteurization), Pasteur studied crystals of tartaric acid and ‘paratartaric acid’ obtained from wine sediments. In particular, he wondered why (as described above) tartaric acid rotated light, while paratartaric acid did not, even though the chemistry and elemental composition of the two were identical. In one of the most beautiful and famous experiments in the history of science, Pasteur noticed, while squinting down a microscope, that there were two subtlety different types of crystals in the samples of paratartaric acid, each the mirror image of the other (see diagram below). He very carefully (and tediously) separated the two types of crystals into separate piles, redissolved each pile, and found that each did indeed rotate light, but in opposite directions. He had, in effect, separated the two enantiomers from the paratartaric acid (a.k.a. racemic acid) and discovered molecular chirality.

The two types of crystals found in paratartaric acid, which are mirror images of each other.

The two types of crystals found in paratartaric acid, which are mirror images of each other.

What does it look like?

The structure of D-tartaric acid (left) and its mirror image, L-tartaric acid (right).

The structure of D-tartaric acid (left) and its mirror image, L-tartaric acid (right).

 

Where did the structure come from?

D-tartaric acid can be found under CCDC refcode TARTAC, while L-tartaric acid is at CCDC refcode TARTAL.

I’m pretty, but don’t touch me as I’m deadly

What does it look like?

Batrachotoxin A from the poison dart frog Phyllobates aurotaenia. Carbon atoms in brown, oxygen atoms in red and nitrogen atoms in blue.

Batrachotoxin A from the poison dart frog Phyllobates aurotaenia. Carbon atoms in brown, oxygen atoms in red and nitrogen atoms in blue.

What is it?

Today is Isabella Karle’s birthday. A leading crystallographer who in her early career worked on the Manhattan project to synthesise plutonium compounds, but became famous for solving the structure of the toxin found in the skin secretions of South American poison dart frogs. Isabella worked with her husband Jerome Karle at the United States Naval Research Laboratory. Jerome Karle won the Nobel Prize for chemistry in 1985 with Herbert Hauptman for their work in X-ray crystallography. Although Isabella didn’t win the Nobel Prize she scored a huge number of awards and prizes in her own right including the US National Medal of Science.

Colombian poison dart frog, Phyllobates aurotaenia. Image from: http://en.wikipedia.org/wiki/Phyllobates_aurotaenia#

Colombian poison dart frog, Phyllobates aurotaenia. Image from: http://en.wikipedia.org/wiki/Phyllobates_aurotaenia#

The toxin shown here is batrachotoxin from the glands of the Dendrobatidae family of frogs found in South and Central America. Native South American tribesmen coat the tips of their hunting darts with the skin secretions of the frogs to give the darts added lethality. The batrachotoxin is extremely lethal requiring only 100 micrograms (or 0.0001 grams) to kill an average sized adult. Batrachotoxin is a neurotoxin and works by binding to the sodium ion channels found in neurons. When batrachotoxin binds to the sodium ion channels the pumps remain permanently open stopping the messages from travelling along neurons and leading to paralysis and heart failure.

Batrachotoxin from frogs consists of three molecules: batrachotoxin A, isobatrachotoxin and pseudobatrachotoxin. The structure shown here is batrachotoxin A that has an additional p-bromobenzoate. The toxin consists of a steroidal structure with its three 6 carbon rings and one 5 carbon ring linked together, and is also an alkaloid due to the presence of the basic nitrogen.

Where did it come from?

The structure was solved by T. Tokuyama and colleagues in 1968 [1] and the structure shown is from the Cambridge Structural Database and published by Isabella Karle in 1969 [2].

[1] Takashi Tokuyama , John Daly , B. Witkop , Isabella L. Karle , J. Karle: The structure of batrachotoxinin A, a novel steroidal alkaloid from the Columbian arrow poison frog, Phyllobates aurotaenia. J. Am. Chem. Soc. (1968) 90, 1917–1918

[2] I.L.Karle, J.Karle: The structural formula and crystal structure of the O-p-bromobenzoate derivative of batrachotoxinin A, C31H38NO6Br, a frog venom and steroidal alkaloid. Acta Cryst. (1969). B25, 428-434

A tree bark suspension and a Nobel Prize later…. We wonder less about the structural basis of the action of aspirin

Today we have another guest post from Lawrence Norris of @BlackPhysicists.  A super read that charts how the every day drug aspirin went from being ancient medicine to how modern crystallography could explain how it works.  Enjoy!

Today’s post spans the bridge between ancient pharmacognosy and modern biophysics. The history of aspirin and its medical use can be traced back to the second millennium BC.  Medicines from willow and other salicylate-rich plants appear in the Egyptian pharmacology papyri.  Hippocrates, the father or modern medicine, administered willow tree bark to women to help relieve pain associated with childbirth.  Later the tree bark was used to alleviate general pain and fevers.

In 1763 Oxford University chemist, Edward Stone, isolated the active compound from Hippocrates preparation, which turned out to be salicylic acid.  In 1853 French chemist Charles Frédéric Gerhardt synthesized acetylsalicylic acid, albeit in an unstable and impure form.  In 1897 German chemist Felix Hoffman, working for Bayer, first synthesize pure acetyl salicylic acid, which later became known as aspirin.  Bayer took aspirin to market almost immediately, and established a complete market association of Bayer AG with the drug.  “Bayer Aspirin” had been on the market for nearly 50 years when Wheatly determined the crystal structure of aspirin in 1964 [1].  There have been several reports of an elusive second crystal polymorphs of aspirin. (See 7 Feb 2014 post on this blog.)

Aspirin-3D-balls.png
Aspirin-3D-balls“. Licensed under Public domain via Wikimedia Commons.

Aspirin took on the moniker of “wonder drug” as it worked wonders against the miseries of aches, pains, fever and inflammation, and scientists wondered why.   Here is where the story of aspirin merges with the Nobel Prize winning work of Bergstrom, Samuelsson and Vane [2] .   This trio and their co-workers elucidated the structure [3] and function of prostaglandins, which are paracrine hormones that mediate many physiological functions having to do with platelet aggregation and smooth muscle tone.   They identified prostaglandin H synthase (PGHS),  as being the key control point enzyme in the “prostaglandin cascade”.   Vane in particular showed that it was possible to block the synthesis and thus the action of prostaglandins with aspirin [4].

Still it was not known exactly how aspirin works.  Enter the work of Picot, Loll and Garavito who solved the crystal structure of PGHS (aka COX)  in 1994 [5].  It bears mentioning here that the crystallization of PGHS, being a membrane protein,  was a tour de force in protein crystallography.  When its structure was published in 1994, there were less than a dozen other membrane proteins in the PDB database.

COX-2 inhibited by Aspirin.png
COX-2 inhibited by Aspirin” by Jeff Dahl – self-made, release to public domain. Licensed under Public domain via Wikimedia Commons.

Following their initial work, Loll, et. al., were able to solve the structure of the enzyme with aspirin bound [6], and also with other drugs that compete with aspirin in the marketplace, e.g., ibuprofen [7].   Others showed how Naproxen (Alleve) binds to the enzyme [8].   Aspirin, it turns out, is an irreversible inhibitor of PGHS.  It acetylates a key serine residue in the enzyme’s active site, and that prevents the natural substrate from situating itself for productive enzyme activity.

Alas though, this is not the whole story of how this wonder drug works physiologically.  Recent results indicate that aspirin is an antiporter partner to protons in mitochondria.  Also aspirin apparently induces formation of inflammation-reducing NO radicals, and signaling through NF-κB.

References
[1]  Wheatley, P. J. “The crystal and molecular structure of aspirin.” J. Chem. Soc., 1163 (1964), 6036-6048, 10.1039/JR9640006036

[2] “The Nobel Prize in Physiology or Medicine 1982”. Nobelprize.org. Nobel Media AB 2014. Web. 25 Nov 2014. <http://www.nobelprize.org/nobel_prizes/medicine/laureates/1982/>

[3]   Abrahamsson, S. “A direct determination of the molecular structure of prostaglandin F2-1.” Acta Crystallographica 16.5 (1963), 409-418, 10.1107/S0365110X63001079

[4]  Vane, J. R. “Inhibition of prostaglandin synthesis as a mechanism of action for aspirin-like drugs.” Nature 231.25 (1971): 232-235, 10.1038/newbio231232a0

[5]  Picot, D, P. J. Loll, and R. M. Garavito. “The X-ray crystal structure of the membrane protein prostaglandin H2 synthase-1.” Nature 367.6460 (1994), 243-249, 10.1038/367243a0

[6]  Loll, P. J., D. Picot, and R. M. Garavito. “The structural basis of aspirin activity inferred from the crystal structure of inactivated prostaglandin H2 synthase.” Nature Structural & Molecular Biology 2.8 (1995), 637-643, 10.1038/nsb0895-637

[7]  Selinsky, Barry S., et al. “Structural analysis of NSAID binding by prostaglandin H2 synthase: time-dependent and time-independent inhibitors elicit identical enzyme conformations.” Biochemistry 40.17,  5172-5180, (2001), 10.1021/bi010045s

[8]  Duggan, Kelsey C., et al. “Molecular basis for cyclooxygenase inhibition by the non-steroidal anti-inflammatory drug naproxen.”  Journal of Biological Chemistry 285.45 (2010), 34950-34959, 10.1074/jbc.M110.162982

 

Crystallising enzymes from beans – celebrating Sumner

What does it look like?

4GY7 jack bean urease deposited in the PDB in 2012. From the paper Crystallographic structure analysis of urease from Jack bean (Canavalia ensiformis) at 1.49 Å resolution deposited by Begum, A.,  Banumathi, S.,  Choudhary, M.I., & Betzel, C

4GY7 jack bean urease deposited in the PDB in 2012. From the paper Crystallographic structure analysis of urease from Jack bean (Canavalia ensiformis) at 1.49 Å resolution deposited by Begum, A., Banumathi, S., Choudhary, M.I., & Betzel, C

What is it and where did the structure come from?

This urease, extracted from a jack bean, was the first enzyme ever to be crystallised meaning that the arrangement of atoms that make it up could then be studied and understood.  The man that achieved this, Jame Batcheller Sumner, was later to win a Noble prize in 1946 for this achievement.  Sunmer, who’s birthday it would be today, actually achieved this feat in 1926 – but it took a number of years to convince the skeptical scientific community that he had isolated an enzyme in his crystals .

James Sumner and his first crystals of an enzyeme

James Sumner and his first crystals of an enzyme

The Nobel Prize in Chemistry 1946 was divided, one half awarded to James Batcheller Sumner “for his discovery that enzymes can be crystallized”, the other half jointly to John Howard Northrop and Wendell Meredith Stanley “for their preparation of enzymes and virus proteins in a pure form”.

Sunmer’s biography on the Nobel Prize website, gives a great insight into how this achievement gradually gained acceptance:

“Gradually, recognition came. In 1937, he was given a Guggenheim Fellowship; he went to Uppsala and worked in the laboratory of Professor The Svedberg for five months. He was awarded the Scheele Medal in Stockholm in the same year. When Northrop, of the Rockefeller Institute, obtained crystalline pepsin, and subsequently other enzymes, it became clear that Sumner had devised a general crystallization method for enzymes. The opponents gradually admitted Sumner’s and Northrop’s claims – Willstätter last of all – and the crowning recognition came in 1946 when the Nobel Prize was awarded to Sumner and Northrop. In 1948, Sumner was elected to the National Academy of Sciences (USA).”

In Sumner’s Nobel acceptance lecture he said:- “Why was it difficult to isolate an enzyme? …i.e. in pure condition. It does not seem difficult to isolate and crystallize an enzyme now, but it was difficult 20 years ago….When I succeeded I then telephoned to my wife ‘I have crystallized the first enzyme.

Tomorrow we’ll have a post about the urease structure in detail, and why it took 85 years from when Sumner first grew the crystals to determining the structure.

References

James B. Sumner The chemical nature of enzymes Nobel Lecture, December 12, 1946

“James B. Sumner – Biographical”. Nobelprize.org. Nobel Media AB 2014. Web. 13 Nov 2014. http://www.nobelprize.org/nobel_prizes/chemistry/laureates/1946/sumner-bio.html

See also:-

P A Karplus, M A Pearson and R P Hausinger 70 Years of Crystaline Urease: What Have We Learned? Acc Chem Res 1997 30, 330-337.

N E Chayen, J R Helliwell and E H Snell “Macromolecular Crystallization and Crystal Perfection” (International Union of Crystallography Monographs on Crystallography” Oxford University Press International Union of Crystallography Monographs on Crystallography (2010).

 

 

Madam Curie – one of her elements Curium

What does it look like?

The double hexagonal close packed structure of Curium.

The double hexagonal close packed structure of Curium.

What is it?

Today would have been Marie Curie’s 147th Birthday.  It’s also medical physics day in honour of the fact that her discoveries lead to the the way we treat many types of cancer with radiation.

Marie Curie’s accolades are well known, amongst other things she was the first person to win both a chemistry and physics Nobel prize.  We’ve actually already covered the curious crystal structure of one of the elements she discovered, Polonium, so instead today we thought we’d cover the element named after her and her husband, Curium.

Curium was first intentionally synthesised in 1944, using a cyclotron at the University of California.  It’s the heaviest of the actinide elements that you can make milligrams of, anything heavier only exists for long enough for a few atoms at a time to exist.  Curium it thought to form a couple of crystal structures, the one we’ve shown here element structure that we haven’t featured on crystallography 365 yet, a double hexagonally closed packed structure. This means that it forms a layered structure, one repeats (the yellow layer in the picture) and in between that two other types of layer (red and blue) form in between that.

Where did the structure come from?

Nobody has ever really made enough curium to do a diffraction study of it, so the structure has been determined from ab initio theory methods by Milman et al in 2003.  You can read more about how they did this in their paper.

Boron – an element to watch for the future

What does it look like?

The arrangement of 12-atom boron clusters in the alpha-tetragonal boron structure.  Image generated by the VESTA (Visualisation for Electronic and STructual analysis) software http://jp-minerals.org/vesta/en/

The arrangement of 12-atom boron clusters in the alpha-tetragonal boron structure. Image generated by the VESTA (Visualisation for Electronic and STructual analysis) software http://jp-minerals.org/vesta/en/

What is it?

It has been a while since we’ve featured an element’s crystal structure on the blog. It’s been said before but we’re continually fascinated that, despite the fact that these crystal structures are made up of just one building block, they can form a whole range of beautiful structures.

Today’s element is no exception to this, boron sits on the top row of the periodic table – it’s the 5th element on the list. One of the features of boron as an element is that it forms clusters with itself. Clusters of 6 atoms, and two arrangement of 12 atoms , and now even a ‘buckyball’ structure.

When you start combining this boron clusters with rare earth elements, then a whole range of interesting properties start to emerge. Many of these materials have been observed to have low thermal conductivity, making them potentially great candidate materials for helping generate thermoelectric power. There’s a wikipedia page that someone has put a lot of crystallographic love into on these materials, in particular scandium and boron complexes.

Where did the structure come from?

We’ve featured one of the first crystal structures of boron, its rare alpha tetragonal allotrope. This was determined in 1951 by Hoard et al, from ‘needle like’ crystals and is reported as a short note in JACS. The structure is #9012288 in the Open Crystallography Database.

Sodium hydride, simple yet historic.

What is it?

Though it looks pretty simple, this structure has a special place in history as it was one of the first structures to be discovered with neutron diffraction.  Though the positions of the sodium atoms in sodium hydride were known for some time (from x-ray diffraction), there was still debate if the hydrogen atoms were at the ½, ½, ½ point in the unit cell (a zinc blende structure) or at the ½, 0, 0 point in the unit cell (a rock salt structure).

In the 1940’s a number of researchers realised that instead of using x-rays to diffract off a material, they could use some different radiation – a beam of neutrons.  This was only possible with the development of nuclear reactors, which could provide a steady source of neutrons.

Neutrons interact with matter in a very different way to x-rays.  With  x-rays, the amount they scatter from an atom is proportional to the number of electrons they have in their outer shell – for instance sodium atoms (with 11 electrons each) scatters much better than hydrogen atoms (with only one electron for each atom).  Instead of interacting with the ‘cloud’ of electrons, neutrons diffract by interacting with the nucleus of the atom itself, and are not affected by it’s size, and so have a much more complex trend as you move through the elements of the periodic table.    For instance heavy hydrogen (deuterium) has a scattering cross length from neutrons that is twice that of sodium.

The effect that is to x-rays the structure of sodium hydride looks like this:

The structure of sodium hydride showing the relative scattering of the sodium (yellow) and hydrogen (pink) atoms.

The structure of sodium hydride showing the relative scattering of the sodium (yellow) and hydrogen (pink) atoms.

and to neutrons it looks like this!

And this shows the relative scattering of the heavy hydrogen (pink) and sodium atoms (yellow) to neutron diffraction.

And this shows the relative scattering of the heavy hydrogen (pink) and sodium atoms (yellow) to neutron diffraction.

This meant that the first researchers using neutron diffraction could work out that the structure of sodium hydride was, in fact, the rock salt structure.  This paved the way for many other structures to be solved with neutron diffraction, locating atoms that would be difficult to see with x-rays.
Shull, one of the authors of this work on sodium hydride, was later to share the Nobel Prize in Physics for his development of neutron diffraction.