E0003

Neutron Protein Crystallography; Case Studies at Current Capabilities and Beyond. Prof. J. R. Helliwell, Dept. of Chemistry, Univ. of Manchester, M13 9PL, UK.

An illustration of the current state of the art and of future needs will be presented using an overview from my own Laboratory's case studies. An example of the state of the art is the hydration structure of the sugar binding protein, concanavalin A, which has been studied by a joint neutron and X-ray protein crystallography approach [1].  This used the Institut Laue Langevin LAue DIffractometer ('LADI').  As an illustration of the need for further neutron facilities development two further case studies will then be presented. Firstly the case of concanavalin A with bound glucoside; this offers very large crystals (100 mm³) but the unit cell size (space group I2₁3, a=168Å) means that the LADI neutron data can be measured to only ~4Å resolution, insufficient for detailed structural analysis [2]. In a final case study, well beyond current neutron facility capability, a description will be given of the structure of the lobster shell colouration protein beta-crustacyanin, which binds astaxanthin.  This structure was determined using SR X-crystallography [3] and involves a bound water molecule at one end of each of the two carotenoids.  The very short H bond distances of the bound water oxygens to the keto oxygens of the astaxanthin end rings (2.65 Å and 2.7Å) arouses an interest in further defining the state of these bound waters especially with respect to a biophysical understanding of the molecular basis of the bathochromic shift effect.  The beta-crustacyanin crystal sample size and the unit cell size puts it completely outside current neutron facility protein crystallographic technical capabilities.  However enhancements of LADI at ILL are planned and, if funded, the proposed European Spallation Source offers further gains.

References.

1. Habash, J., Raftery, J., Nuttall, R., Price, H. J., Wilkinson, C., Kalb (Gilboa), A. J. & Helliwell, J. R. (2000). Direct determination of the positions of the deuterium atoms of the bound water in concanavalin A by neutron Laue crystallography. Acta Cryst. D56, 541-550.

2. Kalb (Gilboa), A. J., Myles, D. A. A., Habash, J., Raftery, J. & Helliwell, J. R. (2001). Neutron Laue diffraction experiments on a large unit cell: concanavalin A complexed with methyl-á-D-glucopyranoside. J. Appl. Cryst. 34, 454-457.

3. Cianci, M., Rizkallah, P J, Olczak, A., Raftery, J., Chayen, N E and Helliwell, J. R. (2002) The molecular basis of the coloration mechanism in lobster shell: beta-crustacyanin at 3.2 Å resolution  PNAS.  USA 99, 9795-9800.