I04-Macromolecular Crystallography
I24-Microfocus Macromolecular Crystallography
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Gustavo
Fenalti
,
Nicolas
Villanueva
,
Mark
Griffith
,
Barbra
Pagarigan
,
Sirish Kaushik
Lakkaraju
,
Richard Y.-C.
Huang
,
Nadia
Ladygina
,
Alok
Sharma
,
David
Mikolon
,
Mahan
Abbasian
,
Jeffrey
Johnson
,
Haralambos
Hadjivassiliou
,
Dan
Zhu
,
Philip P.
Chamberlain
,
Ho
Cho
,
Kandasamy
Hariharan
Open Access
Abstract: CD47 is the only 5-transmembrane (5-TM) spanning receptor of the immune system. Its extracellular domain (ECD) is a cell surface marker of self that binds SIRPα and inhibits macrophage phagocytosis, and cancer immuno-therapy approaches in clinical trials are focused on blocking CD47/SIRPα interaction. We present the crystal structure of full length CD47 bound to the function-blocking antibody B6H12. CD47 ECD is tethered to the TM domain via a six-residue peptide linker (114RVVSWF119) that forms an extended loop (SWF loop), with the fundamental role of inserting the side chains of W118 and F119 into the core of CD47 extracellular loop region (ECLR). Using hydrogen-deuterium exchange and molecular dynamics simulations we show that CD47’s ECLR architecture, comprised of two extracellular loops and the SWF loop, creates a molecular environment stabilizing the ECD for presentation on the cell surface. These findings provide insights into CD47 immune recognition, signaling and therapeutic intervention.
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Sep 2021
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I03-Macromolecular Crystallography
I04-1-Macromolecular Crystallography (fixed wavelength)
I04-Macromolecular Crystallography
I24-Microfocus Macromolecular Crystallography
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Diamond Proposal Number(s):
[12788, 17773]
Open Access
Abstract: Isobutene is a high value gaseous alkene used as fuel additive and a chemical building block. As an alternative to fossil fuel derived isobutene, we here develop a modified mevalonate pathway for the production of isobutene from glucose in vivo. The final step in the pathway consists of the decarboxylation of 3-methylcrotonic acid, catalysed by an evolved ferulic acid decarboxylase (Fdc) enzyme. Fdc belongs to the prFMN-dependent UbiD enzyme family that catalyses reversible decarboxylation of (hetero)aromatic acids or acrylic acids with extended conjugation. Following a screen of an Fdc library for inherent 3-methylcrotonic acid decarboxylase activity, directed evolution yields variants with up to an 80-fold increase in activity. Crystal structures of the evolved variants reveal that changes in the substrate binding pocket are responsible for increased selectivity. Solution and computational studies suggest that isobutene cycloelimination is rate limiting and strictly dependent on presence of the 3-methyl group.
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Sep 2021
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I03-Macromolecular Crystallography
I04-Macromolecular Crystallography
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Diamond Proposal Number(s):
[13775]
Open Access
Abstract: Excessive replication of Saccharomyces cerevisiae Ty1 retrotransposons is regulated by Copy Number Control, a process requiring the p22/p18 protein produced from a sub-genomic transcript initiated within Ty1 GAG. In retrotransposition, Gag performs the capsid functions required for replication and re-integration. To minimize genomic damage, p22/p18 interrupts virus-like particle function by interaction with Gag. Here, we present structural, biophysical and genetic analyses of p18m, a minimal fragment of Gag that restricts transposition. The 2.8 Å crystal structure of p18m reveals an all α-helical protein related to mammalian and insect ARC proteins. p18m retains the capacity to dimerise in solution and the crystal structures reveal two exclusive dimer interfaces. We probe our findings through biophysical analysis of interface mutants as well as Ty1 transposition and p18m restriction in vivo. Our data provide insight into Ty1 Gag structure and suggest how p22/p18 might function in restriction through a blocking-of-assembly mechanism.
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Sep 2021
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Krios III-Titan Krios III at Diamond
Krios IV-Titan Krios IV at Diamond
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Diamond Proposal Number(s):
[20287]
Open Access
Abstract: RNA polymerase inhibition plays an important role in the regulation of transcription in response to environmental changes and in the virus-host relationship. Here we present the high-resolution structures of two such RNAP-inhibitor complexes that provide the structural bases underlying RNAP inhibition in archaea. The Acidianus two-tailed virus encodes the RIP factor that binds inside the DNA-binding channel of RNAP, inhibiting transcription by occlusion of binding sites for nucleic acid and the transcription initiation factor TFB. Infection with the Sulfolobus Turreted Icosahedral Virus induces the expression of the host factor TFS4, which binds in the RNAP funnel similarly to eukaryotic transcript cleavage factors. However, TFS4 allosterically induces a widening of the DNA-binding channel which disrupts trigger loop and bridge helix motifs. Importantly, the conformational changes induced by TFS4 are closely related to inactivated states of RNAP in other domains of life indicating a deep evolutionary conservation of allosteric RNAP inhibition.
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Sep 2021
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E02-JEM ARM 300CF
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Diamond Proposal Number(s):
[16967]
Open Access
Abstract: Single-atom catalysts with maximum metal utilization efficiency show great potential for sustainable catalytic applications and fundamental mechanistic studies. We here provide a convenient molecular tailoring strategy based on graphitic carbon nitride as support for the rational design of single-site and dual-site single-atom catalysts. Catalysts with single Fe sites exhibit impressive oxygen reduction reaction activity with a half-wave potential of 0.89 V vs. RHE. We find that the single Ni sites are favorable to promote the key structural reconstruction into bridging Ni-O-Fe bonds in dual-site NiFe SAC. Meanwhile, the newly formed Ni-O-Fe bonds create spin channels for electron transfer, resulting in a significant improvement of the oxygen evolution reaction activity with an overpotential of 270 mV at 10 mA cm−2. We further reveal that the water oxidation reaction follows a dual-site pathway through the deprotonation of *OH at both Ni and Fe sites, leading to the formation of bridging O2 atop the Ni-O-Fe sites.
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Sep 2021
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Krios II-Titan Krios II at Diamond
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Erika
Chacin
,
Priyanka
Bansal
,
Karl-Uwe
Reusswig
,
Luis M.
Diaz-Santin
,
Pedro
Ortega
,
Petra
Vizjak
,
Belen
Gomez-Gonzalez
,
Felix
Müller-Planitz
,
Andrés
Aguilera
,
Boris
Pfander
,
Alan C. M.
Cheung
,
Christoph F.
Kurat
Diamond Proposal Number(s):
[20287]
Open Access
Abstract: The replication of chromosomes during S phase is critical for cellular and organismal function. Replicative stress can result in genome instability, which is a major driver of cancer. Yet how chromatin is made accessible during eukaryotic DNA synthesis is poorly understood. Here, we report the characterization of a chromatin remodeling enzyme—Yta7—entirely distinct from classical SNF2-ATPase family remodelers. Yta7 is a AAA+ -ATPase that assembles into ~1 MDa hexameric complexes capable of segregating histones from DNA. The Yta7 chromatin segregase promotes chromosome replication both in vivo and in vitro. Biochemical reconstitution experiments using purified proteins revealed that the enzymatic activity of Yta7 is regulated by S phase-forms of Cyclin-Dependent Kinase (S-CDK). S-CDK phosphorylation stimulates ATP hydrolysis by Yta7, promoting nucleosome disassembly and chromatin replication. Our results present a mechanism for how cells orchestrate chromatin dynamics in co-ordination with the cell cycle machinery to promote genome duplication during S phase.
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Sep 2021
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Krios I-Titan Krios I at Diamond
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Diamond Proposal Number(s):
[14393]
Open Access
Abstract: Dynamin belongs to the large GTPase superfamily, and mediates the fission of vesicles during endocytosis. Dynamin molecules are recruited to the neck of budding vesicles to assemble into a helical collar and to constrict the underlying membrane. Two helical forms were observed: the one-start helix in the constricted state and the two-start helix in the super-constricted state. Here we report the cryoEM structure of a super-constricted two-start dynamin 1 filament at 3.74 Å resolution. The two strands are joined by the conserved GTPase dimeric interface. In comparison with the one-start structure, a rotation around Hinge 1 is observed, essential for communicating the chemical power of the GTPase domain and the mechanical force of the Stalk and PH domain onto the underlying membrane. The Stalk interfaces are well conserved and serve as fulcrums for adapting to changing curvatures. Relative to one-start, small rotations per interface accumulate to bring a drastic change in the helical pitch. Elasticity theory rationalizes the diversity of dynamin helical symmetries and suggests corresponding functional significance.
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Sep 2021
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I11-High Resolution Powder Diffraction
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Andrij
Vasylenko
,
Jacinthe
Gamon
,
Benjamin B.
Duff
,
Vladimir V.
Gusev
,
Luke M.
Daniels
,
Marco
Zanella
,
J. Felix
Shin
,
Paul M.
Sharp
,
Alexandra
Morscher
,
Ruiyong
Chen
,
Alex R.
Neale
,
Laurence J.
Hardwick
,
John B.
Claridge
,
Frédéric
Blanc
,
Michael W.
Gaultois
,
Matthew S.
Dyer
,
Matthew J.
Rosseinsky
Diamond Proposal Number(s):
[23666]
Open Access
Abstract: The selection of the elements to combine delimits the possible outcomes of synthetic chemistry because it determines the range of compositions and structures, and thus properties, that can arise. For example, in the solid state, the elemental components of a phase field will determine the likelihood of finding a new crystalline material. Researchers make these choices based on their understanding of chemical structure and bonding. Extensive data are available on those element combinations that produce synthetically isolable materials, but it is difficult to assimilate the scale of this information to guide selection from the diversity of potential new chemistries. Here, we show that unsupervised machine learning captures the complex patterns of similarity between element combinations that afford reported crystalline inorganic materials. This model guides prioritisation of quaternary phase fields containing two anions for synthetic exploration to identify lithium solid electrolytes in a collaborative workflow that leads to the discovery of Li3.3SnS3.3Cl0.7. The interstitial site occupancy combination in this defect stuffed wurtzite enables a low-barrier ion transport pathway in hexagonal close-packing.
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Sep 2021
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I09-Surface and Interface Structural Analysis
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Maximilian
Bauernfeind
,
Jonas
Erhardt
,
Philipp
Eck
,
Pardeep K.
Thakur
,
Judith
Gabel
,
Tien-Lin
Lee
,
Jörg
Schäfer
,
Simon
Moser
,
Domenico
Di Sante
,
Ralph
Claessen
,
Giorgio
Sangiovanni
Diamond Proposal Number(s):
[26419, 25151]
Open Access
Abstract: Large-gap quantum spin Hall insulators are promising materials for room-temperature applications based on Dirac fermions. Key to engineer the topologically non-trivial band ordering and sizable band gaps is strong spin-orbit interaction. Following Kane and Mele’s original suggestion, one approach is to synthesize monolayers of heavy atoms with honeycomb coordination accommodated on templates with hexagonal symmetry. Yet, in the majority of cases, this recipe leads to triangular lattices, typically hosting metals or trivial insulators. Here, we conceive and realize “indenene”, a triangular monolayer of indium on SiC exhibiting non-trivial valley physics driven by local spin-orbit coupling, which prevails over inversion-symmetry breaking terms. By means of tunneling microscopy of the 2D bulk we identify the quantum spin Hall phase of this triangular lattice and unveil how a hidden honeycomb connectivity emerges from interference patterns in Bloch px ± ipy-derived wave functions.
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Sep 2021
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I03-Macromolecular Crystallography
I04-Macromolecular Crystallography
I24-Microfocus Macromolecular Crystallography
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Jiandong
Huo
,
Halina
Mikolajek
,
Audrey
Le Bas
,
Jordan J.
Clark
,
Parul
Sharma
,
Anja
Kipar
,
Joshua
Dormon
,
Chelsea
Norman
,
Miriam
Weckener
,
Daniel K.
Clare
,
Peter J.
Harrison
,
Julia A.
Tree
,
Karen R.
Buttigieg
,
Francisco J.
Salguero
,
Robert
Watson
,
Daniel
Knott
,
Oliver
Carnell
,
Didier
Ngabo
,
Michael J.
Elmore
,
Susan
Fotheringham
,
Adam
Harding
,
Lucile
Moynie
,
Philip N.
Ward
,
Maud
Dumoux
,
Tessa
Prince
,
Yper
Hall
,
Julian A.
Hiscox
,
Andrew
Owen
,
William
James
,
Miles W.
Carroll
,
James P.
Stewart
,
James
Naismith
,
Raymond
Owens
Diamond Proposal Number(s):
[27031]
Open Access
Abstract: SARS-CoV-2 remains a global threat to human health particularly as escape mutants emerge. There is an unmet need for effective treatments against COVID-19 for which neutralizing single domain antibodies (nanobodies) have significant potential. Their small size and stability mean that nanobodies are compatible with respiratory administration. We report four nanobodies (C5, H3, C1, F2) engineered as homotrimers with pmolar affinity for the receptor binding domain (RBD) of the SARS-CoV-2 spike protein. Crystal structures show C5 and H3 overlap the ACE2 epitope, whilst C1 and F2 bind to a different epitope. Cryo Electron Microscopy shows C5 binding results in an all down arrangement of the Spike protein. C1, H3 and C5 all neutralize the Victoria strain, and the highly transmissible Alpha (B.1.1.7 first identified in Kent, UK) strain and C1 also neutralizes the Beta (B.1.35, first identified in South Africa). Administration of C5-trimer via the respiratory route showed potent therapeutic efficacy in the Syrian hamster model of COVID-19 and separately, effective prophylaxis. The molecule was similarly potent by intraperitoneal injection.
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Sep 2021
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