The facilities and staff at the SYBILS beamline contributed to this breakthrough study exploring the extent to which naturally occurring proteins sample the space of folded structures accessible to the polypeptide chain.
Naturally occurring proteins—chains of amino acids that fold into functional, three-dimensional shapes—are believed to represent just a small fraction of the universe of all possible permutations of amino-acid sequences and folds. How can we begin to systematically sift through those permutations to find and engineer from scratch (de novo) proteins with the characteristics desired for medical, environmental, and industrial purposes? To address this question, a team led by researchers from the Institute for Protein Design at the University of Washington have published a landmark study that used both protein crystallography and small-angle x-ray scattering (SAXS) at the ALS to validate the computationally designed structures of novel proteins with repeated motifs. The results show that the protein-folding universe is far larger than realized, opening up a wide array of new possibilities for biomolecular engineering.
Brunette TJ, Parmeggiani F, Huang PS, Bhabha G, Ekiert DC, Tsutakawa SE, Hura GL, Tainer JA, Baker D. “Exploring the repeat protein universe through computational protein design.” Nature 2015 Dec 24 ;528(7583):580-4 PupMed
Our new pixel array detector, a Pilatus3 2M from dectris, arrived at LBNL on July 27, 2015 and was installed on the SAXS endstation at SIBYLS beamline 12.3.1. The first user data was collected on August 19, 2015.
Our 6th annual SIBYLS bioSAXS workshop “Frontiers in biological SAXS” was a huge success. A big thank you to all of you who attended and a special thanks to our fabulous speakers:
Robert Rambo, Principal Beamline Scientist for the solution state SAXS beamline B21 at the Diamond Light Source, Oxfordshire, UK
John J. Tanner, Professor of Biochemistry, University of Missouri-Columbia
Chris Brosey, Postdoc with Tom Ellenberger’s Lab, Washington University in St. Louis
Carrie Partch, Assistant Professor in Physical and Biological Sciences, University of California - Santa Cruz
Samuel Bouyain, Associate Professor, School of Biological Sciences, University of Missouri - Kansas City
Dina Schneidman, Postdoc with Sali Lab, University of California, San Francisco
We look forward to another successful workshop in Fall 2016!
The facilities and staff at the SIBYLS beamline contributed to this recent structural study of a human antibody that is able to neutralize Marburg and Ebola viruses.
The filoviruses, including Marburg and Ebola, express a single glycoprotein on their surface, termed GP, which is responsible for attachment and entry of target cells. Filovirus GPs differ by up to 70% in protein sequence, and no antibodies are yet described that cross-react among them. Here, we present the 3.6 Å crystal structure of Marburg virus GP in complex with a cross-reactive antibody from a human survivor, and a lower resolution structure of the antibody bound to Ebola virus GP. The antibody, MR78, recognizes a GP1 epitope conserved across the filovirus family, which likely represents the binding site of their NPC1 receptor. Indeed, MR78 blocks binding of the essential NPC1 domain C. These structures and additional small-angle X-ray scattering of mucin-containing MARV and EBOV GPs suggest why such antibodies were not previously elicited in studies of Ebola virus, and provide critical templates for development of immunotherapeutics and inhibitors of entry.
Hashiguchi T, Fusco ML, Bornholdt ZA, Lee JE, Flyak AI, Matsuoka R, Kohda D, Yanagi Y, Hammel M, Crowe JE, Saphire EO. “Structural basis for marburg virus neutralization by a cross-reactive human antibody.” Cell 2015 Feb 26 ;160(5):904-12 link
SAXS and MX data collected at SIBYLS enabled insights into protein flexibility in one of the proteins involved in Lou Gerigh’s disease. The results were recently published in PNAS and have been highlighted by Today At Berkeley Lab and and Daily Cal.
Protein framework alterations in heritable Cu, Zn superoxide dismutase (SOD) mutants cause misassembly and aggregation in cells affected by the motor neuron disease ALS. However, the mechanistic relationship between superoxide dismutase 1 (SOD1) mutations and human disease is controversial, with many hypotheses postulated for the propensity of specific SOD mutants to cause ALS. Here, we experimentally identify distinguishing attributes of ALS mutant SOD proteins that correlate with clinical severity by applying solution biophysical techniques to six ALS mutants at human SOD hotspot glycine 93. A small-angle X-ray scattering (SAXS) assay and other structural methods assessed aggregation propensity by defining the size and shape of fibrillar SOD aggregates after mild biochemical perturbations. Inductively coupled plasma MS quantified metal ion binding stoichiometry, and pulsed dipolar ESR spectroscopy evaluated the Cu2+ binding site and defined cross-dimer copper-copper distance distributions. Importantly, we find that copper deficiency in these mutants promotes aggregation in a manner strikingly consistent with their clinical severities. G93 mutants seem to properly incorporate metal ions under physiological conditions when assisted by the cop- per chaperone but release copper under destabilizing conditions more readily than the WT enzyme. Altered intradimer flexibility in ALS mutants may cause differential metal retention and promote distinct aggregation trends observed for mutant proteins in vitro and in ALS patients. Combined biophysical and structural results test and link copper retention to the framework destabilization hypothesis as a unifying general mechanism for both SOD aggregation and ALS disease progression, with implications for disease severity and therapeutic intervention strategies.
Pratt AJ, Shin DS, Merz GE, Rambo RP, Lancaster WA, Dyer KN, Borbat PP, Poole FL, Adams MW, Freed JH, Crane BR, Tainer JA, Getzoff ED. “Aggregation propensities of superoxide dismutase G93 hotspot mutants mirror ALS clinical phenotypes.” Proc. Natl. Acad. Sci. U.S.A. 2014 Oct 14 link
Recently in Science:
Structure determination of gold nanoparticles (AuNPs) is necessary for understanding their physical and chemical properties, but only one AuNP larger than 1 nanometer in diameter [a 102-gold atom NP (Au102NP)] has been solved to atomic resolution. Whereas the Au102NP structure was determined by x-ray crystallography, other large AuNPs have proved refractory to this approach. Here, we report the structure determination of a Au68NP at atomic resolution by aberration-corrected transmission electron microscopy, performed with the use of a minimal electron dose, an approach that should prove applicable to metal NPs in general. The structure of the Au68NP was supported by small-angle x-ray scattering and by comparison of observed infrared absorption spectra with calculations by density functional theory.
read about it here: