Macromolecular Crystallography (MX)

Diamond provides a range of techniques for academic and industrial researchers studying the machines of life. As one of those techniques, Macromolecular Crystallography (MX) reveals the shape and arrangement of biological molecules at atomic resolution, knowledge of which provides a highly accurate insight into function. This can be combined with complementary information from many other techniques available at Diamond alongside lab based investigations to reveal the broader picture of molecular interactions and their effects.

MX is a core activity at Diamond with seven beamlines dedicated to the technique alongside the XFEL Hub, Membrane Protein Laboratory and XChem fragment screening facility for the extensive UK structural biology community as well as researchers in Europe and beyond. The staff of the MX group are recognised as innovative world leaders in MX, moving the goalposts of what is feasible for 'conventional' MX as well as developing techniques and beamlines that transform MX to the next level, enabling new experiments and methodologies. The group takes a long term approach to enabling new capabilities at its suite of beamlines to meet the current and future demands of an exacting community of scientists; in 2017 this was no different, as can be seen by the exciting developments here.
  1. Instruments
  2. Techniques
  3. Compare Beamlines
  4. Pre-Beamtime
  5. Beamtime
  6. Post Beamtime
Instruments -

I02-1 VMXm

Under construction

VMXm is a micro/nanofocus MX beamline aimed at atomic structure determination in cases where the production of significant quantities of protein material and crystals is problematic.

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Detector: Pilatus 6M
Wavelength: 0.57 - 2.0 Å
Energy: 7.0 - 25.0 keV

I02-2 VMXi

Commissioning

The Versatile Macromolecular X-tallography in-situ (VMXi) beamline will be an entirely automated facility for characterisation of, and data collection directly from, crystallisation experiments in situ.

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Beamsize: 10 µm x 10 µm
Energy: 12 keV

I03 MX

High throughput and highly automated beamline for optimised MAD and SAD experiments. Capable of accepting CL3 type experiments on crystals of pathogens.

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Beamsize: variable
Detector: Pilatus3 6M
Wavelength: 0.5 - 2.5 Å

I04 Microfocus MX

Variable focus from 5 to 100 microns, high throughput and highly automated beamline for optimised MAD and SAD experiments.

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Beamsize: 10 µm x 5 µm - 110 µm x 100 µm
Detector: Pilatus 6M-F
Wavelength: 0.69 - 2.07 Å

I04-1 Fixed wavelength MX

High throughput and highly automated fixed wavelength SAD beamline for macromolecular crystallography.

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Beamsize: 60 x 50
Detector: Pilatus 6M-F
Wavelength: 0.92Å (fixed)

I23 Long-Wavelength MX

Commissioning

A unique facility for solving the crystallographic phase problem, using the small anomalous signals from sulphur or phosphorous which are present in native protein or RNA/DNA crystals. Additionally, anomalous difference Fourier maps can be used to locate sulphur and phosphorous positions to assist model building at low resolution and/or identify lighter atoms such as chlorine, potassium and calcium.

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Wavelength: 1.1 – 5.9 Å
Energy: 2.1 – 11 keV

I24 Microfocus MX

High throughput variable microfocus beamline for optimised MAD and SAD experiments on crystals down a few microns in size.

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Beamsize: 5x5 - 50x40 (variable aspect ratio)
Detector: Pilatus3 6M
Wavelength: 0.62Å - 1.77Å

Membrane Protein Laboratory

The Membrane Protein Lab (MPL) at Diamond is a research and training facility for scientists interested in solving the 3D structures of membrane proteins by X-ray crystallography.

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XFEL Hub

The UK is taking a leading role in the development of a new structural biology facility (SFX) at the European X-ray Free Electron Laser (XFEL), in Hamburg, Germany, and a complementary facility at Diamond (The UK XFEL Hub) to help develop the required expertise.

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XChem - Fragment Screening

The fragment screening facility at Diamond Light Source is based in Lab 36 and on beamline I04-1. The scope of the facility is to encompass all stages of the fragment screening experiment.

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Techniques - +

Microfocus

Specialised high flux microfocus beams for microcrystal data collection or the illumination of subvolumes of larger inhomogeneous crystals.

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Remote Access

The highly automated beamlines enable data collection from anywhere in the world via remote access. Systems are in place for shipping samples to and from Diamond Light Source. New users can borrow pucks and tools.

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In situ Data Collection

In situ data collection and sample characterization directly from crystals in their crystallization plates.

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Sample Humidity Control

Control of the sample crystal's relative humidity with the aim of improving diffraction quality through improved packing of protein molecules constituting the lattice.

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Spectroscopy

On-line and off-line UV-Vis spectroscopy of macromolecular samples to study enzyme reaction mechanisms and radiation damage.

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Multi-axis Goniometry

Crystal reorientation to optimize data collection strategies for anomalous diffraction data collection and low symmetry spacegroups, or very long axis unit cells.

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Biocontainment

All beamlines are designated bio containment level 1. For containment level 2 samples beamlines I03 and I24 can be used and beamline I03 can be configured to run at biocontainment level 3.

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Fragment Screening - XChem

The fragment screening facility at Diamond Light Source is based in Lab 36 and on beamline I04-1. The scope of the facility is to encompass all stages of the fragment screening experiment.

More information...
Compare Beamlines - +

 

Beamline
I03 I04 I04-1 I23 I24
Wavelength range (Å)
 0.6 - 2.3 0.69 - 2.07 0.92 (fixed)   0.7 - 2.0
Energy range (keV)
5.2 - 21.0 6.0 - 18.0 13.5 (fixed)   6.4 - 20.0
Default settings (Å/keV) 0.98 / 12.7 0.98 / 12.66 0.92 / 13.53   0.97 / 12.8
Flux (ph/s) in full beam at default energy at 300 mA 1.7 x 1012 2.8 x 1011 9.0 x 1011   3.0 x 1012
Default beamsize (µm) 90 x 20 30 x 20 60 x 50   8 x 8
Full beam size at sample (µm) 90 x 20 90 x 30 100 x100 55 x 10  10 x 5 --> 110 x 100 60 x 50  

5 x 5 --> 50 x 40

Available apertures (µm)

20, 50, 100

  10, 20, 30, 50 and 70    
Detectors (Pilatus type)

P3-6M

(100 Hz)

 P6M-F

(25 Hz)

P6M-F

(25Hz)

Custom P12M

 

P3-6M

(100 Hz)

Multi-Axis Goniometry  Yes Yes Yes Yes Yes
Number of unipucks / SPINE pins 30 / 480 37 / 592 37 / 592   37 / 592
Maximum samples / hour 30 30 30   30
Typical samples / hour 15 - 25  15 - 25 15 - 35   15 - 25 
Pre-Beamtime - +

Plan

Reserve access well ahead of your visit either at the links in the web page or via your local contact to facilities such as

Check the Diamond MX web pages and individual beamline pages for updates, manuals and contact details prior to your experiment.

Register your shipping dewars with Diamond to use our shipping account and help with tracking.

Always purchase and use SPINE pins and Unipucks to mount and ship your samples. Non-SPINE standard pins cause robots to crash and damage to equipment: you lose beamtime, we lose sleep.

For full remote access experiments we prefer pucks to arrive in the shelved shipping cane or alternatively the puck shipping canister. Please avoid the narrow canisters where possible.

Plan

Understand the visit timings you are scheduled for.

For experimenters coming on-site simplify your arrival by taking the Fast Track approach.

For remote experimenters read the remote access guidelines.

  • For remote access experiments the team leader should email their local contact two working days before your visit (by Wednesday afternoon for weekend visits) your users names and contact phone numbers for the visit complete with the number of Dewars and pucks to be loaded. Please use this template to provide this information.
  • For mixed remote access (i.e. where a member of your team is on-site) ensure they will be available to change pucks.
  • In both cases discuss your requirements for remote access with your local contact before your experiment.

For all experiments we highly recommend preparing your shipments in advance in ISPyB, identifying your pucks by the puck identifier (e.g. CPS-0024) and assigning samples at the same time. This allows pucks to be assigned to robot positions as loaded by either you or Diamond staff making them available in GDA immediately. For remote access this is a requirement  rather than a recommendation to ensure reliable information transfer.

For UK users with Diamond registered dewars you can book your next working day shipping to us with Diamond’s DHL account. We recommend shipping your dewars three or more days before your experiment to ensure arrival in time.

Register your PDB and sequences for your proteins in ISPyB. These can then be used automatically if you use default file names in GDA for all future visits in downstream processing such as XIA2, DIMPLE and MrBUMP and avoids the need to do this for each visit.

Beamtime - +

 

Refer to the manual for data collection possibilities –

Read the message of the day in GDA for beamline status.

Refer to beamline specific manuals where appropriate.

Start your data back-up as soon as you can.

Load your pre-registered pucks and assign positions on the touch screen in the hutch (soon in all beamlines). Now start GDA and all your samples will be preloaded in the sample changer view (or press refresh to update if GDA was already running).

Use default file names in GDA to streamline experiments, avoiding the need to type information at each sample load, preventing mistakes and ensuring downstream processing uses the information you have provided in advance (PDB, sequence, space group).

Monitor your results from our automatic software pipelines either in Today's Jpegs webpage, ISPyB or Synchlink.

Talk with your local contact about your experiment plans – we are here to help you solve structures.

Post Beamtime - +

 

Unload your last sample with the sample changer with sufficient time to ensure you have removed pucks from the sample changers before the end of your shift (unless you have discussed a time extension with the local contact).

Deregister your pucks in GDA or on the touch screen in the hutch as you remove them from the sample changer.

Finalise your backup disk and unmount cleanly in the data dispenser interface.

Complete your dewar dispatch form for storage at Diamond or return of your dewars. You can use our shipping agents for dewar return.

Diamond pays for sample shipping (2 dewars per shift) but for UK users only if you use our shipping agents via our webpages. No reimbursement of UK shipping will be made after August 2014 for shipping you arrange yourself.

You can reprocess your data afterwards on our cluster.

Acknowledge beamline access.

 

  • Loan pucks available upon request.
  • New web interface to ISPyB has been launched - please take a look before your visit!
  • New 'Message of the Day' functionality launched in GDA.
  • The SmarGon goniometer installed in I04.
  • The FT-SSX set-up available at I24.
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Beamline Manuals

Open Access

If you have used Diamond MX beamlines remotely via the short-shifts access route anytime after September 2014 or have been a user via BioStruct-X, did you know that your forthcoming peer-reviewed open access publications could be eligible for funding? For more details email openaccess@diamond.ac.uk for beamtime awarded by BioStruct-X in MX/BIOSSAXS or remote.access@diamond.ac.uk to confirm eligibility for remote MX experiments.