REAL-TIME REDUCTION OF AREA DETECTOR DATA BY HARDWARE (DACOM) FOR STATIC AND TIME-RESOLVED CRYSTALLOGRAPHY

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REAL-TIME REDUCTION OF AREA DETECTOR DATA BY HARDWARE (DACOM) FOR STATIC

AND TIME-RESOLVED CRYSTALLOGRAPHY

H. Bartunik, C. Boulin, H. Schwab

To cite this version:

H. Bartunik, C. Boulin, H. Schwab. REAL-TIME REDUCTION OF AREA DETECTOR DATA BY HARDWARE (DACOM) FOR STATIC AND TIME-RESOLVED CRYSTALLOGRAPHY. Journal de Physique Colloques, 1986, 47 (C5), pp.C5-157-C5-166. �10.1051/jphyscol:1986521�. �jpa-00225838�

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Colloque C5, supplement au n o 8, Tome 47, aoQt 1986

REAL-TIME REDUCTION OF AREA DETECTOR DATA BY HARDWARE (DACOM) FOR STATIC AND TIME-RESOLVED CRYSTALLOGRAPHY

H. D

.

'~uropean Molecular Biology Laboratory, c/o DESY, Notkestrasse 85, 0-2000 Hamburg 52, F.R.G.

'"~uropean Molecular Biology Laboratory, Meyerhofstrasse, D-6900 Heidelberg, F.R.G.

. * *

Institut Laue-Langevin, 156 X, F-38042 Grenoble Cedex, France

Abstract - A data handling system ("DACOMn) has been developed for high counting rate (of, at present, 1 MHz and eventually 10 MHz) diffraction data collection with quantum counting or integrating area detectors using synchrotron radiation and for on-line data evaluation in real time. DACOM data handling is done by hardware including look-up tables programmed on the basis of pattern prediction. DACOM may be applied in protein crystallography, both in static studies involving very short exposure and overall measuring times, and in (usec to msec) time-resolved data collection. Results of a test application are described.

I - INTRODUCTION

A number of applications in protein crystallography require both the use of area detectors (ADS) for simultaneous measurement of many reflections and short exposure times as they are achieved with the use of synchrotron radiation (SR). Such applications include in particular crystal structure analysis of protein structures with short lifetimes, due to radiation damage in the X-ray beam, of short-lived intermediates in, e.g.? enzymatic reactions, and rapid (usec-msec) time-resolved cyclic investigation of transient states.

The data rates and the total amount of data to be handled in such applications may be very high. Provided that an adequate AD system is available, a still exposure to high Bragg resolution from a fairly big protein structure like, e.g., Catalase may on a double-focussing beamline on a bending magnet of a storage ring like DORIS be obtained within an exposure time of 10 msec. The exposure time may even be shorter by a factor of lor if a wiggler beamline at DORIS is used. An AD with 512x512 pixels will yield a data rate of about 5 Mbyte/sec, if an exposure time of 100 msec (e.g., for rotation through 0.05g is assumed. Collection

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Article published online by EDP Sciences and available at http://dx.doi.org/10.1051/jphyscol:1986521

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Most of such applications require, in addition to rapid data storage, on-line data analysis of at least part of the data for monitoring the diffraction power of fhe crystal and for dete~ting~changes in crystal orientation or possibly cell dimensions (e.g., in enzyme kinetics).

The possibility to appraise the overall data quality on a similar time scale as the data collection would allow, e.g., to immediately repeat an experiment under modified conditions.

On-line data snalysis on a similar time scale as the SR data collection cannot be realized with software techniques, because of the great number of simultaneously occuring reflections, It is only feasible, if identification of diffraction spots and at least partial integration are done by hardware.

We have developed a system ("DACOMn = data compression) which is capable of handling and reducing data by hardware; its basic concept has been described el~ewhere /I/. DACOM makes use of the fact that the reflections and their positions on the area detector can in many cases be predicted with high accuracy. In the present paper, the detailed concept of DACOM, the DACOM hardware and software, and the results of a test application using experimental AD data are described. The further development of the DACOM system for data rates up to 10 MHz is discussed.

I1 - DATA HANDLING IN HIGH COUNTING RATE APPLICATIONS

The data handling problem in high counting rate applications in protein crystallography using SR may be tackled in basically three alternative approaches:

(1) All original data are transferred via a buffer store onto disk for subsequent evaluation by software. Transfer of 512x512 pixels takes with presently available techniques in the order of 10 sec.

This conventional approach has the advantage that data evaluation may be entirely unbiased and may possibly involve profile fitting /2-4/

leading to optimized peak-to-background. On the other hand, this solution is slow; the data transfer takes by at least a factor of 10 more time than the SR (mini-rotation) exposure. On-line data analysis is not feasible on a time scale which is comparable to that of data collection.

(2) The original data are stored in a buffer, and data are reduced by a multiprocessor system.

Such systems (with typtcally around 4 processors) are being developed in a number of laboratories. The speed of data handling is higher by a maximum factor corresponding to the number of processors. For full on-line analysis, a great number of processors - ideally one per simultaneously observed reflection - would be needed. The systems which are being developed will therefore only rather insignificantly increase the speed of data evaluation.

( 3 ) Information from spot prediction is used to reduce the original

data by hardware, either by transferring the original data only within predicted spot areas, or already by (full or partial) integration over

reflection spots.

Tilis is achieved by the DACOM system which is further described below.

It requires reliable spot prediction. On the other hand, extremely

high speed is obtained, and even time-resolved AD data collection in consecutive time frames is feasible. As a complementary approach, array processors may be used, if the severe problems which appear to still exist both with (the prize of) hardware and software

(D.E.Rimmer, personal communication) can be solved.

I11 - DATA HANDLING BY HARDWARE (DACOM)

The DACOM system is based on the predictability of diffraction patterns for a great number of subsequent exposures. The concept of DACOM and its basic hardware and software are described in the following.

A. Predictability of the Diffraction Pattern

The positional parameters of reflections, including the x,y coordinates on the AD as well as the excitation range in the rotation angle, 4, can in most applications in protein crystallography be predicted, except for a few applications involving, e.g, changes in cell constants during a reaction.

The accuracy in spot prediction may be limited due to a number of effects like crystal slippage or systematic errors in spot prediction.

Crystal slippage has only very rarely been observed in (film) data collection using SR, due to short total measuring times; efficient ADS may speed up data collection even further. Considering possible systematic errors in spot prediction, the shape and the centre positions of a given reflection in x,y and ^f#~ will in general vary as function of the distributions of the mosaic spread of the sample crystal, of the angular divergences and of the wavelengths in the incident beam. The resulting "resolution functionw can in theory be calculated /5/. For protein crystallography, a full calculation of such resolution functions has not yet been carried out; it would be further complicated by the change in the mosaic spread distribution during data collection due to radiation damage. However, the problem has been treated in a practically very useful approximation / 6 / r which has been included in several AD programs as, e.g., the Munich AD program (A.Messerschmidt and J.Pflugrath).

The actual size of such variations in reflection shape and.centre may in SR data collection with ADS be expected to be small, because of the high collimation and the narrow wavelength bandwidth of. the incident beam. It should therefore be possible to achieve high accuracy in spot prediction for a great number of subsequent exposures or even the whole rotation range. However, diagnostic facilities are desired for checking the validity of spot prediction during ongoing data collection.

B. Concept of DACOM

Figure 1 shows a scheme of the basic concept of DACOM fcr the case of.

a quantum counting AD with spatially homogeneous response; the Cases of integrating ADS or heterogeneous response are discussed below. The basic DACOM system consists essentially of a dual-ported 'maskw memory, HM, a self-incrementing "data" store, DM, and a store manager controlling both memories.

Before each exposure, a mask is written into MM which is used as a look-up table. The dimension of the mask cor~esponds to the number of resolution elements of the AD. The mask contains at a given coordinate x,y either Zero ( = no reflection or background spot

C5-160 JOURNAL DE PHYSIQUE

predicted) or a "spot index" characterizing - on the basis of a previous spot prediction

-

I AD READ-OUT I

STORE MANAGER

:KIT

[,,<fi

B F ^{+1 T}

<

C )

X

I

I (T=TIME FRAME)

I

+ I ,."

FURTHER

DISK

'

---

DATA COLLECTIOII DATA COLLECTION DATA COLLECTION

Fig. 1 - Basic concept of DACOM involving a look-up table (mask memory) and a data memory. The flow-chart diagram indicates alternative possibilities for the use of DACOM in static and time-resolved applications and for conventional data collection.

compromise between these extremes; in this mode, a reflection area covering, e.g., 16 pixels may be characterized by, e.g., 4 spot indices. During an exposure, an event with decoded coordinates x,y

initiates the following hardware cycle: The contents (spot index I) of MM at the location x,y is read (without modification1 and presented on an external bus. This spot index I addresses DM for a read-increment-write cycle at location I. After the exposure, DM contains at the location I an integrated countrate. In the case of time-resolved data collection, DM is paged via an external time-frame generator; DM contains eventually at the location IIT an integrated countrate for time frame T. The whole cycle from the AD read-out to incrementation in DM involves only hardware functions; the speed Of data handling is therefore extremely high (see below).

In order to take heterogeneous detector response, varying from pixel to pixel, into account (in case that an integration mapping mode is employed), a second look-up table CM (a "correction" memory) containing detector calibration factors is included in the DACOM system. Further, DM has now to undergo a read-modify-write (rather

than a read-increment-write) cycle.

The hardware cycle is started by

o ~ ~ o o ~ o ~ ~ ~ addressing MM and CM in parallel. o o o o ~ l ~ z o o o o ~ o o ~ DM is addressed at the location

o O E K 4 0 o o o o o o o corresponding to the spot

O ~ O O ~ ~ O ~ O ~ O O index(from MM), and the calibration

o o o o o o LI ~2 ~3 LO o o COLLECT ORIGINAL factor (from CM) is added to the

o o o o o o LS m nz ~ 6o o DATA NlTHlN contents.

0 0 0 0 0 0 L,

, , ,

o o o o o O L ~ N S M ~ U O O o DACOM may also be applied with

o o o o o o u ~ u z u ~ u ~ o o integrating (e.g., TV camera)

~ ~ ~ O ~ O O O O O O O detectors. The system then

~ O ~ O O O O O O O O O includes in addition to MM, CM and DM an arithmetic logical unit (ALU)

O O O O O O O O O O ~ O for hardware multiplication. For

O O K K O O O O O O D O each address (x,y), a countrate N

O O K K O O O O O O O O is presented to DACOM. N is by the

o o o a o o o o o o o o

,,,,

O O O D O O L L L L O O DATA.E.G.,FOR corresponding calibration factor

o o o o o o L o TIME-RES. APPL. (from CM). The product is added to

(INrEGRATION

o o o o o o L n M L o o OVERSPOTS) the contents of DM at the location

O ~ D O O ~ L N ~ L ~ ~ defined by the spot index (from

~ O O O ~ O L L L L O ~ MM)

.

0 0 0 0 0 0 0 0 0 0 0 0

o o ~ o ~ ~ o o o o o ~ In addition to the various mapping modes, the DACOM system allows also Fig. 2

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C. Speed of Data Handling by DACOM

The overall speed of data acquisition-via DACOM is defined by the time needed for

(1) writing a new mask into MM, (2) the hardware operations, and

( 3 ) reading and/or clearing DM.

The rate of data handling during an exposure is only limited by time

( 2 1 , i.e., essentially by the access and cycle times of the memories.

The maximum speed of the hardware used in a first test application of DACOM (see below) was 1.2 MHz. A more advanced version of DACOM, which is in production (C.Boulin), will work at a through-put rate of 10 MHz. Such a data handling rate would allow collection of a full

(900) data set within a total measuring time of about 10 min.

Time (1) is of the order of 1 sec with the present DACOM hardware (see chapter IV). This time can be further reduced

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Time ( 3 ) does practically not limit the total data rate, if DM has two independent pointers; read-out by the computer is then feasible.during

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ongoing data collection (for already fully integrated reflection spots). Clearing DM is in partial or full integration mapping mode anyhow only needed after a great number of exposures or even after completion of the data collection.

D. Complete Reduction of DACOM Data / Diagnostics

As a result of the DACOM hardware cycle, spots are identified through their spot indices, and they are in DM fully or partly integrated according to the mapping mode.

In full integration mapping mode, DM contains after complete excitation of a given reflection a series of partial intensities for each still or rotation exposure through the excitation range in $. ^In

an extreme case, in total only two integrated countrates - ^{for peak}

and background, respectively - are obtained per reflection. In the more general case of several mini-rotations per excitation range, information on the reflection intensity profile is available in a basically similar form as in conventional diffractometer measurements, and full integration and background subtraction may be carried out in essentially the same way. Analysis of the dependence of the partial reflection intensity on provides a basis for diagnostics, e.g., for detecting miscentering in $. Complete data reduction on the basis of partly integrated intensities can be done by the computer in parallel during ongoing data collection.

In partial integration mapping mode, DM contains after full excitation of a given reflection a number of (e.g., 4 ) partial intensities for each exposure. This allows for further diagnostics, in particular for checking the position of the centre in x,y, or for detecting overlap with neighbouring reflections. Otherwise, full data reduction is carried out in a similar way as before, again during ongoing data collection.

In original data mapping mode, DM contains the full AD information for a given reflection within the predicted spot area. Conventional software routines (or possibly an array processor) may be used for centroid determination and full data reduction. This mode may be of interest for specific applications like, e.g., time-resolved studies involving changes in the cell dimensions during a reaction.

E. DACOM Software

A Software package has been developed for data acquisition with DACOM and for complete reduction of DACOM data. The software is compatible with the Munich AD program.

The DACOM software is written in FORTRAN 77 and runs on a VAX 11/750 under VMS. It includes in its present version CAMAC-FORTRAN routines.

The software consists essentially of three components (Figure 3 ) :

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- Data acquisition,

-

This strategy was chosen in order to reduce computer overhead during ongoing data acquisition and to thus achieve minixcum dead time between subsequent exposures.

Preparation of Data Acquisition:

On the basis of a previously determined orientation matrix,

prediction file is created which contains for each reflection the indices HKL, the reflection coordinates on the AD, and the excitation range in the rotation angle. The user defines how to run the experiment by indicating the total rotation range of data collection, static or time resolved data collection mode, the mapping mode etc. A dedicated program scans the range of predicted reflections and assigns locations in DM.

In addition to the prediction file, a mask description file is created for fast update of the mask in MM during the experiment. The mask description file contains for each subsequent exposure the changes to be made relative to the previous mask.

STILt -ures

Refimment parametera

I Prediction

I

I I I

I

r 3

Mm=mQI mom

--

r ^{-t copv} current mask t o M O O W P n D I y

-*+ion crystal me& v a l i d i t y of

s t a r t e x p ~ e u r e

I mt ^for ‘2nd of cxp. preceding e x p ~ s u r r I MapRparenextmak

I

&-Continue Stop

Create data file

Fig. 3 - Flow-chart diagram of the DACOM software.

Data Acquisition:

The acquisition routine controls mainly the crystal rotation (through hardware functions based on beam monitoring) and the update of MM. An image of the current mask is held in the computer core memory; it is vpdated according to the mask description file. Before starting acquisition for the next exposure, the whole MM is overwritten in fast DMA mode. After completion of the data colection through the whole predicted range, the contents of DM is appended to the prediction file and dumped onto disk.

Concurrent Validity Check of Predicted Reflections:

In parallel to the ongoing data acquisition, data in DM which belong to already fully excited reflecti~ns are read and reduced on line. In case that the validity check fails due to, e.g., miscentering in the rotation angle, the acquisition is stopped and, e.g., procedures leading to a refinement of the orientation matrix are anitiated.

IV - TEST APPLICATION OF DACOM

The DACOE software and (the first version off the DACOF: hardware have been tested. In this test application, an experimental protein data set (5O rotation range;, which had previously been collected for Trypsin to 3 fi resolution by A,Messerscbmidt and J.Pflugrath with a

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FAST detector system (ENRAF-NONIUS), was used to simulate data acquisition with an AD. The experimental data were via a CAMAC I/O register presented to the DACOM hardware in essentially the same way as event addresses in real data acquisition with a quantum counter.

CAMAC O A T A W A Y \

SERIAL HIGHWAY TO THE COMPUTER

X TOC Y TDC XY TIME DACOM M A S K DATA SERIAL LRS LRS READOUT R l W E MEMORY HEMORI CRATE SZOl 4201 BEN. SENSION 1623 9.1634 CONTR

DETECTION DIGITISATION COMPRESSION STORAGE INTERFACE

Fig. 4 - ^DACOM hardware (for data rates up to 1.2 MHz) in the context of the complete data acquisition system. For a test application simulating data acquisition, the detection and digitisation parts of the system were replaced by a CAMAC 1/0 register.

E V ~( x y , N Memo?yM& ~ MEMORY PORT ::KglT

0-BUS DUAL

Data CONTROL . ^PORTED

Ready LOGIC

READOUT

frame

= O? LOGIC TI nE FRAME

GENERATOR 1.-

D a t a Taken Ready OM1 i d d r e s s

DATA MCMORY

I

Fig. 5

-

(for data rates

The DACOM hardware (Figure 4) used in the test application consisted of a store manager, a dual-ported Q-bus (mask) memory and a self-incrementing (data) memory. Both (CAMAC) memories with 64K 16-bit were purchased from a commercial company (SENSION). The store manager was developed at the EMBL (C.Boulin). The detailed function of the hardware is schematically described in Figure 5. The CAMAC hardware was controlled by a VAX 11/750 running under VMS.

Mask preparation was done on the basis of a prediction file which was derived from the original spot prediction (A.Messerschmidt and J.Pflugrath) for the Trypsin data. Each reflection spot was characterized

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The partially integrated intensities contained in the data memory were fully integrated and reduced. Background correction was done by using countrates obtained within the same spot area before and after excitation of a reflection.

The structure factors resulting from this DACOM nacquisitionn were merged together using the Munich PROTEIN package. The symmetry R-factor was 7% (in I). This should be compared to an R-factor of 5%

(in I) which was obtained for the same data set by A.Messerschmidt and J.Pflugrath applying the spot centering and .integration routines of the Munich AD program to the complete original data. The difference in the R-factors is essentially explained by the higher peak-to-background ratio which may be obtained with software data evaluation involving dynamical masking /7/.

This test results indicate that even when using DACOM in full integration mapping mode (and hence with limited diagnostics facilities), extremely rapid data collection and on-line data evaluation are indeed possible with an accuqacy, which is comparable to typical results of film data collection.

V

-

The high data rates involved in AD data collection with SR and the importance of on-line analysis of at least part of the data pose severe problems, which may not be solved with conventional techniques based on the storage of original data and subsequent evaluation by software. A solution is, however, feasible through data handling by hardware. This has, for the first time, been achieved with the DACOM system described in this paper; this system may be used both with quantum counting and integrating ADS.

The concept of DACOM is based on the predictability of the positional parameters of the diffraction pattern. This assumption is valid for most applications in protein crystallography. High accuracy in spot prediction has already been achieved with the Munich and a few other AD programs. It may be expected that even higher accuracy may in general be obtained in SR work, because of more favourable resolution functions. Further, the diagnostics facilities included in DACOM allow to check the validity of the current spot prediction on line.

Special mapping modes of DACOM, which involve transfer of original data within predicted spot areas, may be of interest for particular applications, e.g., in cases when changes in cell dimensions may occur during data collection.

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DACOM may be used in static high-speed protein data collection with, at present, data rates of more than 1 MHz; this has been demonstrated in a test application using experimental data. A more advanced version of DACOM, which is in production, will handle data rates of 10 MHz. However, the use of DACOM may also be of interest in medium counting rate applications with data rates of a few 100 kHz for on-line analysis of data in parallel to conventional storage of the original data.

In addition to static work, DACOM provides for the first time a means for rapid (usec-msec) time-resolved AD data collection; this is also of potential importance for other types of diffraction studies like, e.g., time-resolved investigation of contracting muscle.

Data collection with DACOM (in full integration mapping mode) provides basically a similar type of information as single counter diffractometer measurements. DACOM data may be of similarly high quality, if the intrinsic AD properties like signal-to-noise and time stability are adequate. As compared to AD data collected with conventional techniques, DACOM data will be less affected by radiation damage to the sample, because of much shorter total measuring times.

DACOM may be further developed at reasonable costs for even higher data rates, more detector resolution elements or a greater number of simultaneously excited reflections. The dimensions of memories, which constitute the fundamental components of the DACOM hardware, may be extended (e.g., for multiplexing) at relatively low (and decreasing) prizes. In total, it may be expected that AD data handling by hardware will play a more and more important role in high counting rate applications.

REFERENCES

/1/ Bartunik,H.D. and Boulin,C., in Structural Biological Applications of X-Ray Absorption, Scattering and Diffraction, Eds.

H.D.Bartunik and B.Chance, Acad. Press, N.Y. 1985 (in press).

/2/ Ford,G.C., J, Appl. Cryst. 7 (1974) 555.

/3/ Kabsch,W., J. Appl. Cryst. 10 (1977) 426.

/4/ Rossmann,M.G., J. Appl. Cryst. 12 (1979) 225.

/5/ Cooper,M.J. and Nathans,R., Acta Cryst. A24 (1968) 619.

/6/ Greenhough,T.J. and Helli~el1,J.R.~ J. Appl. Cryst. 15 (1982) 338.

/7/ Sjglin,L.G. and Wlodawer,A., Acta Cryst. A37 (1981) 594.

ACKNOWLEDGMENT

A.Messerschmidt and J.Pflugrath supported this work by making experimental AD data and the corresponding spot prediction file available to us for a test application of DACOM; we wish to express our gratitude for their most valuable help and for many discussions.

We would further like to thank S.MacLaughlin for her assistance in part of the programming. Helpful discussions with M.Boehm and C.Rieke1 are acknowledged.