K. Sakabe

Weissenberg camera for macromolecules with imaging plate data collection system at the Photon Factory: Present status and future plan (invited)

A Weissenberg camera for macromolecules with imaging plate data collection system at BL6A and BL18B stations in the Photon Factory is introduced and evaluated. The special feature of these systems is considered matching for both SR‐x rays and protein crystallography. This system is user‐friendly and can collect a large amount of data to higher resolution from the crystal with large unit cell dimensions. A newly developed camera can be used as a time‐resolved Laue camera and a Weissenberg camera. A large image reader (IPR4080) that can scan 400×400 mm2 and 400×800 mm2 sizes of imaging plates has been developed and evaluated. The new data collection system combined with the new camera and IPR4080 will be installed at the BL18B station in October 1994.

Extracellular nucleases of Bacillus subtilis I. Purification and properties

Abstract Four Ca2+-dependent nucleases, nucleases Bs-10, Bs-IA, Bs-IB and Bs-II, have been purified from a culture fluid of Bacillus subtilis SB19 and their properties investigated. Nucleases Bs-IA, Bs-IB and Bs-II hydrolyze native and denatured DNA and rRNA in the presence of Ca2+. Without the addition of Ca2+, denatured DNA and rRNA are degraded at reduced rates and little degradation of native DNA is found. The activities of these enzymes are inhibited by EDTA and other agents which bind Ca2+. The mode of action of these enzymes on DNA is predominantly exonucleolytic and the major products from DNA are deoxyribonucleoside 3′-monophosphates. However, an endonuclease activity at the level of one-hundredth of the gross nucleolytic activity is detected in the purified preparations of these enzymes. The enzyme nuclease Bs-10 is different from the other three enzymes in that it hydrolyses only denatured DNA and only to a limited extent.

X-ray studies of water structure in 2 Zn insulin crystal

The water structure of rhombohedral 2 Zn insulin crystal which contains about 280 water molecules and 0.55-0.60 mol citrate molecules per dimer has been studied by X-ray crystallographic refinement with 1.1 A resolution data. Atomic parameters of 83 fully occupied and 258 partially occupied water molecules and 0.3 mol of citrate were obtained. Full matrix least-squares method with isotropic temperature factor was used for the refinement of partially occupied water molecules. The water molecules in this crystal exist in one of the three states: fully occupied water, partially occupied water and water continuum, and a schematic model of water structure in protein crystal was proposed. The flexibility of water molecules is described.

Highly bright X-ray generator using heat of fusion with a specially designed rotating anticathode.

A very compact X-ray generator, 4.3 times more brilliant than can be attained by a conventional rotating-anticathode X-ray generator, has been developed using a U-shaped rotating anticathode and a high-flux electron gun with focusing bending magnet.

Synchrotron radiation protein data collection system using the newly developed Weissenberg camera and imaging plate for crystal structure analysis (abstract)

It has been an earnest desire of protein crystallographers to collect fast, accurate, high resolution diffraction data from protein crystals, preferably with exposure time as short as possible. In order to meet this challenge, a new type of Weissenberg camera has been developed for the recording of diffraction intensity from protein crystals using synchrotron radiation. The BL6A2 line has a plane‐bending mirror designed by Y. Sato. The optical bench with triangular tilt‐cut Si crystal monochromator was designed by N. Kamiya and was installed in the BL6A2 hutch. The Weissenberg camera was set on the 2θ arm of the optical bench. This camera can be used with Fuji Imaging Plate (IP) as an x‐ray detector, and the reading out of the image from the IP is carried out by using BA100. The characterization of this system was carried out using the native crystal of chicken gizzard G‐actin DNase I complex and its Yb3+, PCMB, indium, and FMA derivatives. Since these crystals are very sensitive for x rays, the resolutio...

Time-Resolved Protein Crystallography with Large-Angle Oscillations: an Application of a Protein Data-Collection System Using the Weissenberg Technique and a Large-Format Imaging Plate

A diffraction-intensity data-collection system with synchrotron radiation X-rays utilizing the screenless Weissenberg technique and incorporating a large-format imaging plate is one of the most suitable apparatus for time-resolved protein crystallography with larger angle oscillations than hitherto described. The time resolution and data quality of the system have been tested using a tetragonal lysozyme crystal as a test sample in a flow-cell experiment at the bending-magnet beamline 18B at the Photon Factory, and a time resolution of 15 min is confirmed.

Conceptual design of novel IP-conveyor-belt Weissenberg-mode data-collection system with multi-readers for macromolecular crystallography. A comparison between Galaxy and Super Galaxy.

Galaxy is a Weissenberg-type high-speed high-resolution and highly accurate fully automatic data-collection system using two cylindrical IP-cassettes each with a radius of 400 mm and a width of 450 mm. It was originally developed for static three-dimensional analysis using X-ray diffraction and was installed on bending-magnet beamline BL6C at the Photon Factory. It was found, however, that Galaxy was also very useful for time-resolved protein crystallography on a time scale of minutes. This has prompted us to design a new IP-conveyor-belt Weissenberg-mode data-collection system called Super Galaxy for time-resolved crystallography with improved time and crystallographic resolution over that achievable with Galaxy. Super Galaxy was designed with a half-cylinder-shaped cassette with a radius of 420 mm and a width of 690 mm. Using 1.0 A incident X-rays, these dimensions correspond to a maximum resolutions of 0.71 A in the vertical direction and 1.58 A in the horizontal. Upper and lower screens can be used to set the frame size of the recorded image. This function is useful not only to reduce the frame exchange time but also to save disk space on the data server. The use of an IP-conveyor-belt and many IP-readers make Super Galaxy well suited for time-resolved, monochromatic X-ray crystallography at a very intense third-generation SR beamline. Here, Galaxy and a conceptual design for Super Galaxy are described, and their suitability for use as data-collection systems for macromolecular time-resolved monochromatic X-ray crystallography are compared.

Large format IP scanner with multi-reading heads for automated protein crystallography using SR

Foundation for Advancement of International Science,SBSP House, KEKPF,1-1 Oho,Tsukuba,Ibaraki 305-0801,Japan Rigaku Corporation,3-912,Matsubara,Akishima,Tokyo196-8666,Japan Macscience. Corporation,78 Nagaya,Sakai-cho,Sakai-gun,Fukui 919-0598,Japan KEKPF,Oho,Tsukuba,Ibaraki 305-0801,Japan Division of Biological Sciences, Graduate School of Science, Hokkaido University,Kita-10, Nishi-8, Kita-ku, Sapporo 060-0810, Japan Graduate School of Environmental Studies Nagoya University,Furo-cho, Chikusa, Nagoya 464-8601, Japan