Photon Factory Activity Report 2005 #23 Part B (2006)
Biological Science

10A/2005G161

X-ray diffuse scattering from protein crystals caused by the lattice defects.
Takuya ISHIKAWA1, Daichi YAGI1, Toshikatsu ICHIGE1, Yuusuke SAWA1, Kenji KAWAMURA1,Yuuki OHNISHI2, Ichiro TANAKA3, Izumi YOSHIZAKI4, Nobuo NIIMURA1
1Graduate School of Science and Engeneering, and 3College of Engeneering, Ibaraki Univ. Hitachi,
24316-8511, Japan, Kaken Co. Ltd., Mito, Japan and JAXA, Tsukuba, Japan
Introduction
Recent high resolution (better than 1Å) X-ray proteincrystallographic analysis requires high quality protein
crystals. High quality crystals in turn produce a highresolution diffraction data set. In protein crystallography
the quality of the crystal is usually defined as follows: anoften-used alternative criterion is whether the crystal
produces a high resolution Bragg reflection data set ornot. One possible explanation of the quality of protein
crystals is whether the protein molecules areorientationally ordered within the unit cell. In that case,the overall B-factor obtained by the Wilson plottechnique [1] becomes a good measure of the quality ofthe protein crystals. As a matter of the fact, we havealready made such a proposal and we have proved thatthe overall B-factors obtained from Wilson plots correlatemuch better with the quality of the crystals
experimentally [1]. In other words, the quality of theprotein crystals is discussed by the degree of the disorderof the protein molecules in the unit cell.
Generally in X-ray diffraction phenomena it is wellknown that the disorder of molecules in unit cells
provides diffuse scattering under a Bragg reflection
profile. In the past such a diffuse scattering from a protein
crystal has been often observed and this phenomenon has
been discussed from the view point of moleculardynamics. We do not deny that one part of the diffuse
scattering is the contribution of molecular dynamics, but
we think that the main part is one of the disorder ofmolecules, incompleteness of crystals. The diffraction
pattern on the imaging plate observed at 77K shows very
distinctive diffuse scattering under Bragg reflections,which do not depend on temperature. We tried to measure
the accurate profile of diffuse scattering in order toanalyze the relation between the crystal quality and themolecular disorder.

Experiment
We have carried out the preliminary measurement of
the accurate profile of diffuse scattering in order toanalyze the relation between the crystal quality and themolecular disorder.
We have used the 4 circle diffractometer at BL10A in
PF of KEK. The wavelength is 1.54 Å Figure 1 shows therocking curve profile of the Bragg reflection of cubic
insulin. The FWHM is 6/1000 deg. A diffuse scattering isseen under the Bragg reflection from the logarithmic
display of the profile as shown in Fig.2. We will continue
the measurement of the diffuse scattering from several
protein crystals as a function of crystallization conditionand crystal quality determined by B factors in order todiscuss the nature of the quality of protein crystals.
90000

80000

70000

60000

50000

40000

30000

20000

10000

0
9.57

9.58

9.59

9.6

9.61

9.62

9.63

Fig.1 Rocking curve profile of the Bragg reflection.
14
12
10
8
6
4
2
0-0.001-0.0008-0.0006-0.0004

0-0.0002

0.00020.00040.00060.00080.001-2-4-6Sample Preparation
Porcine insulin was purchased from Wako Pure
Chemicals Industries, Ltd. Insulin readily dissolves inpure water with the addition of ammonium hydroxide.
Crystals of cubic insulin were grown by a dialysis method
at several crystallization condition such as at different
protein and precipitant concentrations. (The precipitant isNa2HPO4.)

2

Fig.2 Ln I vs. Q display of Fig.1.
Reference[1] S.Arai, T.Chatake, N.Suzuki, H.Mizuno,N.Niimura,Acta Cryst. D60, 1032-1039 (2004)
niimura@mx.ibaraki.ac.jp

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