Newsletter contents... UP

CCLRC Daresbury Laboratory

Protein Crystallography Specialist Users Group Meeting

York University, Thursday 4 January 2001

Report by Pierre Rizkallah

Agenda changes by the SR Users Forum at the SRS Users Meeting in September 2000 meant that the usual Specialist Users sessions to cover individual scientific areas were not convened. However, users of the Protein Crystallography facilities on the SRS expressed an interest in holding an alternative Specialist Users Group meeting at a convenient time and place. An appropriate opportunity presented itself in the form of a satellite meeting prior to the CCP4 Study Weekend, normally held early in January. The CCP4 organisers were very kind to extend support and assistance for the SUG meeting, and it was held the day before the start of the Study weekend. The delegates arriving early at York appreciated the opportunity for discussing common topics of interest. There were some 35 delegates, representing many of the user groups from the UK.


A scientific presentation from Robert Esnouf, Oxford University, discussed the usage of anomalous diffraction from S atoms, in the structure determination of a salivary protein from ticks. Although there had been high-resolution data available, collected at SRS Station 9.6 to a resolution of 0.9 Å, the structure could not be solved with direct methods. Data were then collected on SRS Station 14.1 at a wavelength of 1.488 Å, where the f’’ signal from S atoms was equal to almost 0.5 e‾. Such a small signal was countered by a high redundancy data set recorded over 855°. The resolution was curtailed due to the geometry limitations, but I/sig(I) was around 30 in the outermost shell, 1.7 - 1.6 Å. Shake’n’Bake located 16 S atoms in the 8 disulphide bridges of the 2 protein molecules in the asymmetric unit, using the Bijvoet differences. Phase extension to higher resolution was successful with ACORN. Chain tracing with ARP/wARP located all the ordered residues. The final stages of the refinement used SHELX. Further map interpretation will be needed to model the mobile N-termini.

Michele Cianci described the usage of longer wavelength (2.0 Å) in the structure determination of a-crustacyanin, a lipocalin. The similar protein structures available in the PDB could not be used in Molecular Replacement. The softer X-rays were used to collect a native data set with 20-fold redundancy, a data set with Xe derivative, and another native data set under similar conditions to those of the Xe derivative. All these data sets were limited by diffraction geometry and detector dimensions to a resolution of 2.3 Å. Finally, a 1.2 Å data set was collected with high intensity short-wavelength X-rays (0.87 Å). The Xe anomalous data set was used in Shake’n’Bake to solve the Xe structure. The 4 sites found were used to determine the positions of the S atoms. The correct hand was determined by verification against the high redundancy data set. The phases were used in the high-resolution data set and extended to cover all the resolution range. The various density manipulation techniques were used to get a good quality map, which was traced automatically with WARPnTRACE. All the sequence could be traced and minimal intervention was needed. The final model was refined against data to 1.35 Å resolution, with an R-factor of 17.5% and an R-free of 22.2%. Comparison with the other lipocalins is now underway.

Colin Nave then reported on the status of the SRS. After the vacuum leak in Nov 2000, which forced an early shutdown, repairs have been completed successfully, and the RF system upgraded, with a good prognosis for a smooth restart in January 2001. There were longer-term plans for wiggler cryogenics replacement and a further upgrade of the RF system. An extra degree of flexibility has been introduced into the overall scheduling system by incorporating "contingency days" which will either be shutdown or scheduled beam depending on previous events. Colin also reminded those present of the possibility of requesting extra or alternative days by sending an e-mail message to, after inspecting the current schedule at New ionising radiation regulation requires users to be instructed in the safe usage of each station, and a form to be filled logging this training. A member of staff would be available at the weekend to provide this training for groups starting on weekend days, and to assist with incidental problems. Planned developments of the PX facilities included robotics and automatic systems for sample changing, centring, data collection and processing. Remote experiment control by operators at their home institution will also be developed. All the main data analysis software was made available at the stations, so users can complete their experiment while still at DL. A new development is also underway to build a new multipole wiggler beamline, in Straight 10, over the next two years. This will be Beamline 10, for MAD applications.

The station managers then reported on their respective stations:

The final part of the meeting was a general discussion session where participants addressed many topics:

  1. Facility developments should continue apace, in a ‘run up’ to DIAMOND. Methodology should be made immediately transferable when DIAMOND comes on-line
  2. Station improvements are continuing, and feedback was called for, to ensure optimal response to the users needs.
  3. The Biology Programme Joint Steering Committee have approved funding for automation developments, specifically a sample changer.
  4. The grant for Beamline 10 would be administered by the Research Councils, and the beamline was due for completion earl in 2003. The construction work was forecast to impact on Line 9, causing some station closures for short periods.
  5. A stream of projects should be built up to capitalise on the new facilities being developed, especially in the wake of automation.
  6. Flexible scheduling is under evaluation. Extensive usage was asked for, in order to investigate potential shortcomings.
  7. The participants encouraged the development of a longer wavelength data collection facility, either on Station 7.2 or Station 9.5. A rapid CCD system would be necessary for the collection of the required high redundancy data.
  8. Automation was envisaged as an important factor for crystal screening, remote monitoring of experiments, and, in due course, remote control of experiments by scientists at their laboratory.
  9. A PX service was under consideration, to offer operator data collection for qualifying users.

The participants expressed satisfaction at the alternative arrangements made for the Specialist Users Group session. They also expressed their gratitude to the CCP4 organisers for accommodating this session at short notice, and would hope to make it a regular feature in the coming years.

Newsletter contents... UP