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LADI - Protein crystals

A series of short movies showing how neutron experiments are prepared or performed. Some of these movies have an historical interest since they captured important moments in the life of the ILL.

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Films and animations


Protein crystals from space for neutron studies

LADI is a Laue diffractometer LADI mainly used for single-crystal studies of small protein systems at medium or high resolution in order to locate individual hydrogen atoms of special interest, water structures or other small molecules that can be marked with deuterium to be particularly visible.

Since neutron diffraction requires larger sample than X-ray diffraction, growing large enough protein crystals is thus a prerequisite. Crystallization in microgravitation is one amongst techniques which have emerged in that field. The movies below show Flora Meilleur (EMBL-Grenoble, then ILL) performing a preparatory test with cytochrome p450 in normal gravitation, a red metalo-protein (the color is due to its iron contents).
The crystallization cell shown here is similar to those shipped by NASA on the Spacelab. How does it work ? The precipitating agent (PEG8000) diffuses within the protein solution through a dialysis membrane which results in the crystallization of the protein in the solution. PEG8000 also diffuses from 2nd to the 1st chamber (through an agarose gel) and this continuous input of precipitating agent within the protein solution results in the growth of larger crystals.

Movie by Alain Filhol.

©2004-2007 Institut Laue-Langevin, Grenoble, France.

Movie files (288x352 pixels, duration: 3'45"): .mp4 (7.1 Mb), .webm (23 Mb)
Movie files with no subtitle (576x768 pixels, duration 1'00"): .mp4 (17.1 Mb)

Sequences of the movie

English version:
1- Showing the cell and its various parts;
2- Flora preparing the cell;
3- Doses of cytochrome p450 recently prepared at EMBL-Grenoble;
4- Flora taking a small quantity of protein and placing it into one of the recipients of the cell caps;
5- Removing a membrane from the beaker, which is then placed on the protein container;
6- Removing the excess membrane;
7- Putting the lid on the cell;
8- Filling one of the chambers with a precipitating agent solution (PEG8000);
9- Putting the sealing cap on the chamber;
10- The cell is turned upside down;
11- Filling the second chamber with a more concentrated precipitating agent solution (PEG8000);
12- Putting the sealing cap on the second chamber;
13- Removing excess solvent... The cell now is ready!
14- The cell seen from above with the protein (red) well visible through the central window;
15- A few hours later, the protein has crystallized, but under the form of a poly-crystal. Therefore, tests will have to continue!

French version:
1- Présentation de la cellule et de ses divers éléments
2- Flora prépare la cellule
3- Doses de cytochrome p450 récemment préparées à l'EMBL
4- Flora prélève une petite quantité de protéine et la place dans le réceptacle de l'un des capuchons de la cellule
5- Prélèvement d'une membrane dans un Becher. Elle est ensuite placée sur le réceptacle de la protéine et est maintenue par un joint
6- Découpage de la partie de la membrane qui dépasse
7- Mise place du capuchon sur le corps de la cellule
8- Remplissage de la première chambre avec une solution d'agent précipitant (PEG8000)
9- Mise en place du bouchon de fermeture de la première chambre
10- La cellule est retournée
11- Remplissage de la seconde chambre avec une seconde solution d'agent précipitant (PEG8000) plus concentrée
12- Mise en place du bouchon de fermeture de la seconde chambre
13- Elimination du solvant en excès... La cellule est prête !
14- La cellule vue de dessus avec la protéine (rouge) bien visible à travers la fenêtre centrale
15- Quelques heures plus tard, la protéine a cristallisé mais sous forme polycristalline. D'autres tests seront donc nécessaires.

English checked by: Ronen Ghosh, April 2007.
Updates: A.Filhol, 17 Sept 2008, 8 Sept 2009, 16 April 2012.

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