R. Hamlin

The electron stationary picture method for high-speed measurement of reflection intensities from crystals with large unit cells

A method for measuring X-ray intensity data from crystals with large unit ceils is presented. The method takes full advantage of the capabilities of the multiwire area detector diffractometer. This diffractometer is a high-speed data collection system utilizing a multiwire proportional chamber that can detect photons from all simultaneously occurring reflections and record the diffraction pattern as a two-dimensional histogram in a computer mass core memory. In the electronic stationary picture method of data collection, reflection intensities are extracted from a sequence of electronic pictures each of which is exposed while the crystal is stationary. Between successive pictures the crystal is rotated about a fixed axis by a small constant angle of approximately 0.05°. Because the integrated intensity of each reflection is extracted from several consecutive pictures, advance prediction of detector coordinates and picture number is required for all reflections and the data-extraction computer program is necessarily complex. Expressions are derived for reflection detector coordinates and setting angles, the system hardware and software are described, the data collection procedure is outlined, and the quality of 7 × 105 reflection intensities measured during six months of operation is analyzed.

A multiwire proportional chamber as an area detector for protein crystallography

A multiwire proportional chamber (30 × 30 cm) together with its electronic readout into a large core memory (mass core) has been used successfully as a digital area detector for protein crystallography. The diffraction pattern stored in the mass core can be displayed on a TV monitor. An IBM 1800 computer has fast random access to the mass core and is used on line to estimate the integrated reflection intensities. To characterize this new area detector, the geometric linearity, the resolution and the quantum detection uniformity have been measured. Preliminary results show that with this new system one can collect intensity data from protein crystals about an order of magnitude faster than with the standard diffractometer.

Characteristics of a flat multiwire area detector for protein crystallography

A flat, relatively thin (9 mm) xenon-filled multiwire proportional counter with two-dimensional, 2/~s delay line readout of a 270 x 300 mm active area has been developed for use as a position-sensitive area X-ray detector in the 8 keV energy region (Cu Ka) used in crystallographic structure work with large biological molecules. Its quantum detection efficiency for 8 keV Xray photons is about 0.5, a value which is spatially uniform to within _+ 2%. Its dead-time loss fraction at a typical data collection rate of 30 000 photons s-1 is 12%. The detector has spatial resolution for X-rays of 0.6 mm FWHM in the horizontal direction and 2 ram, the anode wire spacing, in the vertical direction. The effects of parallax are found to be limited and do not seriously increase the apparent size of the diffracted beams. The position sensitivity of this detector is geometrically linear to within 0-5 mm across its active surface. Routine maintenance of the detector requires the attention of a skilled technician but is not time consuming. For four years, this detector has been used to measure millions of reflection intensities from crystals of many different proteins. The down time due to the detector has averaged less than four days per year, considerably less than the down time of other components of the data collection system. Four new protein structures have now been solved using data from this detector. Also, a considerable amount of data have been collected at higher resolution or at different temperatures with crystals of other proteins.

Structure determination of histidine decarboxylase from Lactobacillus 30a at 3.0 A resolution.

The crystal structure of histidine decarboxylase from Lactobacillus 30a has been determined by X-ray diffraction methods to a resolution of 3.0 A. This protein is a pyruvoyl-dependent enzyme that is formed by an unusual self-activation process. The structure was determined from an electron density map calculated using multiple isomorphous replacement phases from two heavy-atom derivatives and included contributions from anomalous scattering measurements. The final mean figure of merit was 0.79, based on 28,805 independent reflections. The molecule has an (alpha beta)6 subunit composition and crystallizes in the space group 14122 with a = b = 221.7 A and c = 107.1 A. There is one (alpha beta)3 half molecule per asymmetric unit. The (alpha beta)6 particle is dumbbell-shaped, with each (alpha beta)3 unit being approximately spherical, with a diameter of about 65 A. There is a large central cavity approximately 30 A deep around the molecular 3-fold axis of the (alpha beta)3 unit. The 3-fold related active site pockets are located around the bottom of this cavity and are separated from each other by a distance of approximately 23 A. The inner portion of each (alpha beta) unit, which lies near the interface between the two (alpha beta)3 particles, consists mainly of random coil with several small helical and sheet regions. The outer region of each (alpha beta) unit has an unusual structure consisting of two overlapping, predominantly antiparallel beta-pleated sheets, lined on each side by an alpha-helix. The walls of the central cavity are formed by the 3-fold repeat of two strands from this beta-sandwich structure and one of the helices.

Progress report on the development of an area detector data acquisition system for X-ray crystallography and other X-ray diffraction experiments

Abstract A relatively inexpensive data acquisition system, incorporating a flat MWPC, with an anticipated count rate of approximately 130 kHz is currently being developed at SSRL in collaboration with research groups at UCSD and LBL. It will be primarily used for X-ray crystallographic studies of macromolecules using synchrotron radiation but will also be used for other X-ray diffraction experiments at SSRL. This system, controlled by a PDP 11-34 computer running under the RSX-11M operating system, incorporates the MWPC area detector, a fast display system, a 3-circle single-crystal goniometer, an alignment carriage, an incident-beam intensity monitor and a tunable monochromator. A progress report and future plans will be presented. The detector dimensions are 28 cm × 28 cm with a spatial resolution of 128 × 256 elements. The absorption depth at normal incidence is 1 cm and the fill gas is a mixture of 90% Xe and 10% CO 2 at a pressure of 1 atmosphere. Fast printed-circuit delay-lines (≈ 170 ns) are used for read-out. It is anticipated that the complete system will be ready for initial testing in the Spring of 1980.

A xenon-filled multiwire arc detector for X-ray diffraction

A xenon-filled multiwire proportional chamber with high and uniform photon-detection efficiency has been built and incorporated into a high-speed data-collection system for protein crystallography. This system has been used to collect more than 200 000 reflection intensities from one parent and four heavy-atom derivative crystals of a complex of dihydrofolate reductase and methotrexate (a cancer drug). Its data collection rate is about 30 times faster than the rate of a standard diffractometer (1400 reflection intensities per hour versus 50 reflection intensities per hour). The precision of the chamber data is also better with an intensity reliability R of 5% as compared with 6.7% from the diffractometer data. The difference Patterson maps between parent and heavy-atom derivative data show very clearly the positions of the heavy atoms.

Development of a Mixed-Mode Pixel Array Detector for macromolecular crystallography

A Mixed-Mode Pixel Array Detector (MMPAD) has been conceived for protein crystallography applications. The most notable feature of the detector is the combination of analog and digital approaches used in the pixel to generate very large dynamic range, low dead time, and fast readout of the types of spot intensity measurements needed in X-ray images from large macromolecules. Prototype structures for the pixel have been fabricated and evaluated and indicate that an intensity range of 10/sup 7/ to 1 can be measured with a precision of 0.25%, while still maintaining a high DQE for X-rays in the 6-18 keV range. The mixed-mode structure is compatible with a fast I/O system and allows the possibility of full-frame readout in the ms range. We present details of pixel behavior, design of support electronics, hybridization results and plans for fabrication of large arrays.

An area detector data acquisition system for X-ray crystallographic studies of macromolecules and other X-ray diffraction experiments☆

Abstract A data acquisition system, incorporating a flat multi-wire proportional chamber, is currently being developed at SSRL in collaboration with research groups at UCSD and LBL [1]. It will be primarily used to rapidly measure X-ray crystallographic data at multiple wavelengths for the investigation of anomalous scattering techniques in the field of protein crystallography. This system, which is in the final stages of development, is controlled by a PDP11/34 computer and incorporates the MWPC area detector, a Huber 5-circle single-crystal goniometer, an alignment carriage, an incident-beam intensity monitor and a fast television display system. An outline of the complete system and the proposed method of data collection will be presented.