I03-Macromolecular Crystallography
I04-1-Macromolecular Crystallography (fixed wavelength)
I04-Macromolecular Crystallography
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Diamond Proposal Number(s):
[10369, 14692]
Open Access
Abstract: The bacterial second messenger cyclic-di-GMP is a widespread, prominent effector of lifestyle change. An example of this occurs in the predatory bacterium Bdellovibrio bacteriovorus, which cycles between free-living and intraperiplasmic phases after entering (and killing) another bacterium. The initiation of prey invasion is governed by DgcB (GGDEF enzyme) that produces cyclic-di-GMP in response to an unknown stimulus. Here, we report the structure of DgcB, and demonstrate that the GGDEF and sensory forkhead-associated (FHA) domains form an asymmetric dimer. Our structures indicate that the FHA domain is a consensus phosphopeptide sensor, and that the ligand for activation is surprisingly derived from the N-terminal region of DgcB itself. We confirm this hypothesis by determining the structure of a FHA:phosphopeptide complex, from which we design a constitutively-active mutant (confirmed via enzyme assays). Our results provide an understanding of the stimulus driving DgcB-mediated prey invasion and detail a unique mechanism of GGDEF enzyme regulation.
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Sep 2019
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I04-1-Macromolecular Crystallography (fixed wavelength)
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Jon A.
Read
,
Jonathan
Tart
,
Philip B.
Rawlins
,
Clare
Gregson
,
Karen
Jones
,
Ning
Gao
,
Xiahui
Zhu
,
Ron
Tomlinson
,
Erin
Code
,
Tony
Cheung
,
Huawei
Chen
,
Sameer P.
Kawatkar
,
Andy
Bloecher
,
Sharan
Bagal
,
Daniel H.
O’donovan
,
James
Robinson
Abstract: Enhancer of zeste homologue 2 (EZH2), the catalytic subunit of polycomb repressive complex 2 (PRC2), regulates chromatin state and gene expression by methylating histone H3 lysine 27. EZH2 is overexpressed or mutated in various hematological malignancies and solid cancers. Our previous efforts to identify inhibitors of PRC2 methyltransferase activity by high-throughput screening (HTS) resulted in large numbers of false positives and thus a significant hit deconvolution challenge. More recently, others have reported compounds that bind to another PRC2 core subunit, EED, and allosterically inhibit EZH2 activity. This mechanism is particularly appealing as it appears to retain potency in cell lines that have acquired resistance to orthosteric EZH2 inhibition. By designing a fluorescence polarization probe based on the reported EED binding compounds, we were able to quickly and cleanly re-triage our previously challenging HTS hit list and identify novel allosteric PRC2 inhibitors.
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Sep 2019
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Open Access
Abstract: A new copper(I) pyrazine-bridged coordination polymer [Cu2(pyz)3(NO3)2]·2DMF] (pyz = pyrazine) (1) has been synthesized and characterized by FT-IR, TG/DTG, DSC and single crystal X-ray diffraction techniques. The X-ray crystallographic result reveals a two-dimensional network structure containing hexagonal pores. Thermal analysis of compound 1 reveals it is stable to 380 °C, and gas sorption studies showed that it adsorbs 1.04 wt% hydrogen at 1 atm and 77 K. Compound 1 crystallizes in a triclinic system, space group P-1 (no. 2), a = 7.9550(2) Å, b = 7.9810(2) Å, c = 11.0660(3) Å, α = 76.328(1)°, β = 71.115(1)°, γ = 84.577(1)°, V = 645.79(3) Å3, Z = 2, T = 150(2) K, μ(MoKα) = 1.709 mm-1, Dcalc = 1.639 g/cm3, 11111 reflections measured (7° ≤ 2Θ ≤ 54.96°), 2951 unique (Rint = 0.0539) which were used in all calculations. The final R1 was 0.0346 (>2σ(I)) and wR2 was 0.0727 (all data).
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Sep 2019
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I04-Macromolecular Crystallography
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Diamond Proposal Number(s):
[17212]
Abstract: Neprilysin (NEP) is a promiscuous zinc metalloprotease with broad substrate specificity and cleaves a remarkable diversity of substrates through endopeptidase action. Two of these – amyloid‐β and natriuretic peptides – implicate the enzyme in both Alzheimer’s disease and cardiovascular disease, respectively. Here, we report the creation of a catalytically inactive NEP (E584D) to determine the first peptide‐bound crystal structure at 2.6 Å resolution. The structure reveals key interactions involved in substrate binding which we have identified to be conserved in other known zinc metalloproteases. In addition, the structure provides evidence for a potential exosite within the central cavity that may play a critical role in substrate positioning. Together, these results contribute to our understanding of the molecular function of NEP.
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Sep 2019
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I02-Macromolecular Crystallography
I03-Macromolecular Crystallography
I04-1-Macromolecular Crystallography (fixed wavelength)
I04-Macromolecular Crystallography
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Rosalba
Lepore
,
Andriy
Kryshtafovych
,
Markus
Alahuhta
,
Harshul A.
Veraszto
,
Yannick J.
Bomble
,
Joshua C.
Bufton
,
Alex N.
Bullock
,
Cody
Caba
,
Hongnan
Cao
,
Owen R.
Davies
,
Ambroise
Desfosses
,
Matthew
Dunne
,
Krzysztof
Fidelis
,
Celia W.
Goulding
,
Manickam
Gurusaran
,
Irina
Gutsche
,
Christopher J.
Harding
,
Marcus D.
Hartmann
,
Christopher S.
Hayes
,
Andrzej
Joachimiak
,
Petr G.
Leiman
,
Peter
Loppnau
,
Andrew L.
Lovering
,
Vladimir V.
Lunin
,
Karolina
Michalska
,
Ignacio
Mir‐sanchis
,
Alok
Mitra
,
John
Moult
,
George N.
Phillips Jr
,
Daniel
Pinkas
,
Phoebe A.
Rice
,
Yufeng
Tong
,
Maya
Topf
,
Jonathan D.
Walton
,
Torsten
Schwede
Diamond Proposal Number(s):
[14692]
Open Access
Abstract: The functional and biological significance of selected CASP13 targets are described by the authors of the structures. The structural biologists discuss the most interesting structural features of the target proteins and assess whether these features were correctly reproduced in the predictions submitted to the CASP13 experiment.
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Sep 2019
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I02-Macromolecular Crystallography
I03-Macromolecular Crystallography
I04-Macromolecular Crystallography
I24-Microfocus Macromolecular Crystallography
Krios I-Titan Krios I at Diamond
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Diamond Proposal Number(s):
[19432, 18659, 12579]
Open Access
Abstract: Plants, algae, and cyanobacteria fix carbon dioxide to organic carbon with the Calvin–Benson (CB) cycle. Phosphoribulokinase (PRK) and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) are essential CB-cycle enzymes that control substrate availability for the carboxylation enzyme Rubisco. PRK consumes ATP to produce the Rubisco substrate ribulose bisphosphate (RuBP). GAPDH catalyzes the reduction step of the CB cycle with NADPH to produce the sugar glyceraldehyde 3-phosphate (GAP), which is used for regeneration of RuBP and is the main exit point of the cycle. GAPDH and PRK are coregulated by the redox state of a conditionally disordered protein CP12, which forms a ternary complex with both enzymes. However, the structural basis of CB-cycle regulation by CP12 is unknown. Here, we show how CP12 modulates the activity of both GAPDH and PRK. Using thermophilic cyanobacterial homologs, we solve crystal structures of GAPDH with different cofactors and CP12 bound, and the ternary GAPDH-CP12-PRK complex by electron cryo-microscopy, we reveal that formation of the N-terminal disulfide preorders CP12 prior to binding the PRK active site, which is resolved in complex with CP12. We find that CP12 binding to GAPDH influences substrate accessibility of all GAPDH active sites in the binary and ternary inhibited complexes. Our structural and biochemical data explain how CP12 integrates responses from both redox state and nicotinamide dinucleotide availability to regulate carbon fixation.
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Sep 2019
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I05-ARPES
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R. C.
Vidal
,
H.
Bentmann
,
T. R. F.
Peixoto
,
A.
Zeugner
,
S.
Moser
,
C.-h.
Min
,
S.
Schatz
,
K.
Kissner
,
M.
Unzelmann
,
C. I.
Fornari
,
H. B.
Vasili
,
M.
Valvidares
,
K.
Sakamoto
,
D.
Mondal
,
J.
Fujii
,
I.
Vobornik
,
S.
Jung
,
C.
Cacho
,
T. K.
Kim
,
R. J.
Koch
,
C.
Jozwiak
,
A.
Bostwick
,
J. D.
Denlinger
,
E.
Rotenberg
,
J.
Buck
,
M.
Hoesch
,
F.
Diekmann
,
S.
Rohlf
,
M.
Kalläne
,
K.
Rossnagel
,
M. M.
Otrokov
,
E. V.
Chulkov
,
M.
Ruck
,
A.
Isaeva
,
F.
Reinert
Diamond Proposal Number(s):
[19278, 22468]
Abstract: The layered van der Waals antiferromagnet
MnBi
2
Te
4
has been predicted to combine the band ordering of archetypical topological insulators such as
Bi
2
Te
3
with the magnetism of Mn, making this material a viable candidate for the realization of various magnetic topological states. We have systematically investigated the surface electronic structure of
MnBi
2
Te
4
(0001) single crystals by use of spin- and angle-resolved photoelectron spectroscopy experiments. In line with theoretical predictions, the results reveal a surface state in the bulk band gap and they provide evidence for the influence of exchange interaction and spin-orbit coupling on the surface electronic structure.
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Sep 2019
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I16-Materials and Magnetism
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C. T.
Koops
,
S. B.
Hrkac
,
M.
Abes
,
P.
Jordt
,
J.
Stettner
,
A.
Petraru
,
H.
Kohlstedt
,
V.
Hrkac
,
N.
Wolff
,
L.
Kienle
,
O. H.
Seeck
,
G.
Nisbet
,
O. M.
Magnussen
,
B. M.
Murphy
Abstract: The elastic coupling in multiferroic materials and even more so in magnetoelectric composites plays an important role for the properties and function. In this study, the electrically induced strain at the epitaxial interface of 0.72Pb(Mn1/3Nb2/3)O3–0.28PbTiO3—CoFe2O4, a magnetoelectric composite, is characterized by in situ X-ray scattering experiment and transmission electron microscopy study. For the measured range all strain induced lattice changes are reversible ruling out plastic deformation. The surprisingly non-perfect elastic coupling of 87 ± 7% in this epitaxial system can be explained by the presence of planar defects in the CFO film.
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Sep 2019
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I10-Beamline for Advanced Dichroism
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Abstract: B2-ordered FeRh undergoes a first-order metamagnetic transition from an antiferromagnet (AF) to a ferromagnet (FM) upon heating. Thin films of B2-ordered FeRh are grown using DC magnetron sputtering and characterized for their behaviour at GHz frequencies and their dynamic behaviour over hour timescales. Ferromagnetic resonance investigations reveal a change in the spectroscopic splitting factor, g, through the range of the transition probed here. By introducing a model that describes the development of the two magnetic phases through the transition, this change in g is shown to be consistent with the development of an exchange coupling across the magnetic phase boundary that induces a non-zero magnetic moment in the AF phase as the result of a thickness dependent phase transition in the AF layer. The influence of such a phase transition is also seen in the extracted value of the Gilbert damping parameter in this experiment. Spin-wave resonance measurements are then performed to try and conclusively measure the exchange coupling between the two magnetic states in FeRh. Measuring the exchange stiffness through the transition reveals that the AF phase has a non-zero exchange energy that varies through the measurement range probed here. The behaviour of the exchange stiffness in the AF layer is attributed to a combination of both the onset of the exchange coupling and the presence of evanescent spin-waves, both of which are consequences of the thickness dependent phase transition in the AF layer. It was then shown that the structure of both magnetic phases could be measured directly with X-Ray Magnetic Dichroism using both linearly and circularly polarized light. The objects measured in these experiments are then characterized for their dynamic properties using X-Ray Photon Correlation Spectroscopy. These studies reveal reveal that the dynamic behaviour of the system is dependent on the type of magnetic dichroism used to probe it. This study also shows it is possible use x-ray magnetic linear dichroism to directly measure the structural and dynamic behaviour of AF materials.
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Sep 2019
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I03-Macromolecular Crystallography
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Diamond Proposal Number(s):
[1984]
Abstract: Computational design of disulfide bonds was performed for lipase from Geobacillus stearothermophilus T6 (LipT6) for enhanced methanol stability and improved biodiesel production. Thirteen double mutants comprising new cysteine pairs were screened and evaluated for their stability in 70% methanol. Superior stability was found with variant E251C/ G332C (M13) having a 5.5‐fold higher hydrolysis activity and enhanced unfolding temperature (Tm) of +7.9°C in methanol compared with wild‐type. Moreover, M13 converted nearly 80% waste chicken oil to biodiesel, representing a 2.4‐fold improvement relative to the WT. Structural studies using X‐ray crystallography confirmed the existence of the engineered disulfide bonds shedding light on the link between the bond location and backbone architecture with its stabilization impact. Rational integration of disulfide bonds is suggested to be a feasible method to promote elevated stability in organic solvents for various industrial applications such as biodiesel synthesis.
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Sep 2019
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