Celebrating Laue – comprehensive protein structures

In the last of our three posts celebrating Max von Laue and Laue diffraction, Prof John Helliwell tells us how the technique has impacted on protein crystallography.

What is it?

To get a comprehensive picture of a protein structure in its functional state(s) can lead to seeking a time-resolved ‘movie’. But the X-ray data collection may be too slow to capture the interesting moment of function. As an alternative to use of monochromatic radiation the use of polychromatic X-ray synchrotron radiation leads to much quicker diffraction data collection. Moreover the sought after hydrogenation and bound water details may be incomplete as hydrogen scatters X-rays weakly. But this latter challenge can be overcome by use of neutron diffraction, as deuterium scatters neutrons as strongly as e.g. carbon. But neutron beams are much weaker in intensity than synchrotron X-ray beams.

As in the X-ray case above though the polychromatic neutron emission from either a nuclear reactor or a pulsed source can be more efficiently harnessed using Laue diffraction and again leads to shorter exposure times. Research in the 1980s [1,2] led to a much improved understanding of Laue crystallography and greatly assisted accurate Laue diffraction data analysis along with newly developed software.

Concanavalin A is isolated from jack beans; approx 10% by weight of protein is con A. Its role is not known for sure but is implicated in an anti fungal protection strategy for the bean via protein to protein  cross linking involving polysaccharide.

Concanavalin A is isolated from jack beans; approx 10% by weight of protein is con A. Its role is not known for sure but is implicated in an anti fungal protection strategy for the bean via protein to protein
cross linking involving polysaccharide.

Overall this has led to a variety of time-resolved Laue X-ray protein crystallography and neutron Laue crystallography studies [eg 3,4]. As just one example the crystal shown here, ~3mm long, is of the lectin protein concanavalin A, (isolated from jack beans ~1cm, also pictured), and was used both to help develop these modern Laue methods and to reveal new details of its bound water structure at the sugar molecular recognition binding site both at room temperature and at 15K [4].

 

What does the structure look like?

Concanavalin A is a tetramer of 100 kDa

A crystal of jack bean concanavalin A,  saccharide free crystal form.

In a final twist of this story the neutron Laue study of concanavalin A at 15K [4] showed that freeze quench time-resolved studies with neutrons even of large crystals of a protein was possible. In the ten years since this study [4] there have been various further improvements in neutron beam fluxes, measuring apparatus and use of fully deuterated proteins in Europe, Japan and the USA radically changing the scope of neutron macromolecular crystallography to smaller crystals and larger molecular weights of proteins.

[1] D.W.J. Cruickshank, J.R. Helliwell and K. Moffat ‘Multiplicity Distribution of Reflections in Laue Diffraction’. Acta. Cryst. A43, 656-674 (1987).

[2] D.W.J. Cruickshank, J.R. Helliwell and K. Moffat ‘Angular distribution of reflections in Laue diffraction’ Acta Cryst. (1991) A47, 352–373.

[3] J.R. Helliwell, Y.P. Nieh, J. Raftery, A. Cassetta, J. Habash, P.D. Carr, T. Ursby, M. Wulff, A.W. Thompson, A.C. Niemann and A. Hädener “Time–resolved structures of hydroxymethylbilane synthase (Lys59Gln mutant) as it is loaded with substrate in the crystal determined by Laue diffraction” (1998) Faraday Trans. 94(17), 2615–2622.

[4] M.P. Blakeley, A.J. Kalb (Gilboa), J.R. Helliwell & D.A.A. Myles (2004) “The 15-K neutron structure of saccharide free concanavalin A” Proceedings of the National Academy of Sciences USA 101, 16405-16410. PDB code 1XQN.

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Celebrating Laue – very tiny crystals.

Yesterday we introduced Laue diffraction, and discussed how it is useful for rapid data collection.  Today’s application is a bit different – what if you’ve only got a very small crystal?

What does it look like?

Image generated by the VESTA (Visualisation for Electronic and STructual analysis) software http://jp-minerals.org/vesta/en/

Image generated by the VESTA (Visualisation for Electronic and STructual analysis) software http://jp-minerals.org/vesta/en/

 

What is it?

With the advent of more powerful synchrotron sources, this is less of a problem than it used to be.  But when you only had a very tiny crystal (say about 20 microns, the size of a cell found in your glands) is was very difficult to get good enough diffraction data to determine what they were.  So in 1995 a group looked at using Laue diffraction to tell between tiny crystals of minerals with similar structures.  These were cerussite (lead carbonate) and strontianite (strontium carbonate).  As they have very similar structures, the researchers had to examine the intensity of the diffraction peaks to tell if each tiny crystal had lead or strontium in it.

Where did the structure come from?

The structure of cerussite was originally determined by Colby and LaCoste in 1933, and is #1010956 in the open crystallographic database.  

Celebrating Laue – complex structures very quickly!

In a slight oversight at crystallography 365 HQ, we missed that it was Max von Laue’s birthday on Thursday.

von_laueVon Laue received the Nobel prize for physics in 1914 for his discovery of the diffraction of x-rays by crystals, the first crucial step to the science of crystallography as it is today.  Though von Laue went on to concentrate on theoretical physics, one particular technique of diffraction is named after him – Laue diffraction.   With this technique of diffraction the sample is kept still and the x-rays it is exposed to are polychromatic – they occur in a range of wavelengths.  This means that lots of planes of atoms satisfy the Bragg condition at once, and makes for some very pretty pictures.

An image of Laue diffraction from http://staff.chess.cornell.edu/~hao/research.html

An image of Laue diffraction from http://staff.chess.cornell.edu/~hao/research.html

Laue diffraction is particularly useful for a number branches of crystallography – and over the next few days we’ll be featuring a few of them.  Today we’ll introduce one use – taking rapid data!

What does it look like?

Image generated by the VESTA (Visualisation for Electronic and STructual analysis) software http://jp-minerals.org/vesta/en/

Image generated by the VESTA (Visualisation for Electronic and STructual analysis) software http://jp-minerals.org/vesta/en/

What is it?

As you can see this is a pretty complex arrangement of atoms, it’s a complex of the metal rhodium in a state that only exists for a few seconds at a time.  The advantage of Laue diffraction, is that if you can push the material into its excited state as well as hitting it with a pulse of x-rays – you can find even the most fleeting of crystal structures.

Where did the structure come from?
This is structure came from work by Makal et al. and is #2019360 in the open crystallographic database.