ADSC was very helpful in making some subtle adjustments to the detector gain maps so that the background of the images collected with our newly upgraded Q315r detector are much more uniform.
Before After
The "Before" image is a 2 second exposure with the multilayer optics and a fully tuned beam. In addition we had some plastic cover slips in the beam to generate background scatter. The "After" image is also a 2 seconds, but with the Si(111) optics and no plastic in the beam path. We will confirm that the gain remap has fully fixed the checker syndrome, but the initial images look very promising.
An excellent paper came out today in the Oct 3rd issue of Cell detailing structural, biochemical, and genetic studies of the Mre11-DNA complex and its role in detecting and repairing double-strand breaks in DNA. Both the SAXS and crystallography data were collected at the SIBYLS beamline. There is also a nice writeup by Paul Preuss which appears in the todays Berkeley Labs News Release.
“Mre11 dimers coordinate DNA end-bridging and nuclease processing in double-strand break repair” by R. Scott Williams, Gabriel Moncalian, Jessica S. Williams, Yoshiki Yamada, Oliver Limbo, David S. Shin, Lynda M. Groocock, Dana Cahill, Chiharu Hitomi, Grant Guenther, Davide Moiani, James P. Carney, Paul Russell, and John A. Tainer, appears in the 3 October 2008 issue of Cell.
The SIBYLS beamline Kohzu monochromator has often exhibited hysteresis when moving the theta 2 motor. This manifests itself in a decaying beam and suboptimal intensity. In order to fully understand the problem it helps to have a general understanding of the motors in the monochromator. The figure below illustrates the various motors and their relative motions.
The axis of the Theta motor lies on the surface of the first crystal and the angle of Theta determines the wavelength of X-rays that will be selected for a particular experiment. The second crystal is designed to take that monochromatic X-ray beam coming from the first crystal and reflect it down the beam pipe to the SAXS or MX endstation. The second crystal is ~30 meters from the crystal sample when doing a crystallography experiment and it is critical that the um-sized X-ray beam be aimed directly at the small crystalline samples. We use the Chi 2 motor on the second crystal to steer the beam from left to right (horizontal beam steering) and we use the M2 mirror to move the beam up and down (vertical beam steering). The axis of the Theta 2 motor lies on the surface of the second crystal, and it is used to align the surface of the second crystal so that it is perfectly parallel to the first crystal thus maximizing the flux of the X-rays that exits the monochromator. Typically we tuneup the beamline before a user starts their shift and during this tuneup procedure we optimize the angle of the theta 2 motor by monitoring an ion gauge positioned at the exit tube of the mono (Imono Out) and maximize the current. If we then monitor the Imono Out gauge for several hours after the tuneup procedure is complete we sometimes see that the intensity of X-rays falls precipitously.
In the September 19, 2008 issue of Cell, we report striking conformational rearrangements in the crystal structure of NEDD8~Cul5ctd-Rbx1 and SAXS analysis of NEDD8~Cul1ctd-Rbx1 relative to their unmodified counterparts. These results point to conformational control of Cullin-RING ligase (CRL) activity, with ligation of NEDD8 shifting equilibria to disfavor inactive closed architectures, and favor dynamic, open forms that promote polyubiquitination.
Cullin-RING ligases (CRLs) comprise the largest ubiquitin E3 subclass, in which a central cullin subunit links a substrate-binding adaptor with an E2-binding RING. Covalent attachment of the ubiquitin-like protein NEDD8 to a conserved C-terminal domain (ctd) lysine stimulates CRL ubiquitination activity and prevents binding of the inhibitor CAND1. Here we report striking conformational rearrangements in the crystal structure of NEDD8~Cul5ctd-Rbx1 and SAXS analysis of NEDD8~Cul1ctd-Rbx1 relative to their unmodified counterparts. In NEDD8ylated CRL structures, the cullin WHB and Rbx1 RING subdomains are dramatically reoriented, eliminating a CAND1-binding site and imparting multiple potential catalytic geometries to an associated E2. Biochemical analyses indicate that the structural malleability is important for both CRL NEDD8ylation and subsequent ubiquitination activities. Thus, our results point to a conformational control of CRL activity, with ligation of NEDD8 shifting equilibria to disfavor inactive CAND1-bound closed architectures, and favor dynamic, open forms that promote polyubiquitination.
Due to DOE budget cuts and planned upgrades to the ALS storage ring we will have no light until October 11th.
Hopefully we will be scheduling beamtime for Oct/Nov/Dec within the next couple of weeks. If you are interested in obtaining General Users shifts please fill out an online application.