Krios IV-Titan Krios IV at Diamond
|
Diamond Proposal Number(s):
[25452, 32707]
Open Access
Abstract: Antimicrobial resistance is driving the search for new antibiotics and a greater understanding of their mechanism of action. Doxycycline is amongst the most-prescribed antimicrobials. It demonstrates a particularly low minimum inhibitory concentration against the zoonotic pathogen Coxiella burnetii. Doxycycline canonically targets the bacterial ribosome by blocking tRNA binding at the decoding centre (A site) of the small subunit. Using cryo-electron microscopy, we analysed doxycycline binding to C. burnetii and Escherichia coli ribosomes. Both structures reveal doxycycline binding at the exit tunnel in the large subunit. In C. burnetii three doxycycline molecules stack to block the tunnel. In E. coli one doxycycline molecule triggers a major change in the conformation of the ribosome. This rearrangement of the peptidyl transferase centre blocks tRNA binding and nascent chain accommodation, abolishing interactions that are fundamental to ribosome function. We identify a distinct ribosomal protein in the C. burnetii large subunit and characterise an additional member of the prokaryotic ribosome hibernation-promoting factor family. These insights into ribosome function and antibiotic action may aid the development of new ribosome inhibitor antibiotics.
|
Jun 2026
|
|
Krios III-Titan Krios III at Diamond
Krios IV-Titan Krios IV at Diamond
|
Douglas S. M.
Ferreira
,
Katie
Mclennan
,
Calum
Diamond
,
Melanie
Vollmar
,
Wasim
Kiyani
,
D. Sean
Froese
,
Jola
Kopec
,
Henry J.
Bailey
,
Rod
Chalk
,
Arnaud
Basle
,
Jonathan M.
Elkins
,
Jesse
Coker
,
Wyatt W.
Yue
,
Thomas J.
Mccorvie
Diamond Proposal Number(s):
[34172]
Open Access
Abstract: Human methionine synthase (MTR) is an essential enzyme of one carbon metabolism. Consisting of a catalytic N-half and a cobalamin binding C-half, MTR utilises this intricate organometallic cofactor in the methyl transfer from methyltetrahydrofolate to homocysteine producing methionine. Cobalamin loading into MTR, and its subsequent activation, requires methylmalonic aciduria and homocystinuria Type D (MMADHC) protein and methionine synthase reductase (MTRR), respectively. However, the molecular basis of cobalamin binding and activation of human MTR aided by MMADHC and MTRR remains unknown. Here, using cryo-electron microscopy, we determine structures of human MTR in its apo, and cobalamin bound states. Apo MTR adopts a conformation where the two halves of the enzyme act independently with the C-half posed to bind cobalamin. Binding of cobalamin and its activation causes conformational changes in MTR that result in a flexible catalytically active state. AlphaFold predictions, validated by interaction studies, show that MMADHC interacts with the C-half of apo MTR to facilitate cobalamin loading. Unexpectedly we found that MTRR interacts at two distinct sites within the C-half of MTR which may aid in activation. Collectively these findings lay the groundwork to uncover the mechanisms through how MMADHC and MTRR coordinate cobalamin loading and activation of human MTR.
|
May 2026
|
|
Krios I-Titan Krios I at Diamond
Krios II-Titan Krios II at Diamond
Krios IV-Titan Krios IV at Diamond
|
Diamond Proposal Number(s):
[29812, 34631, 21005]
Open Access
Abstract: RNA polymerase II (RNA Pol II) is central to gene expression, but its catalytic mechanism remains elusive due to the absence of high-resolution structural data. The role of water molecules in RNA Pol II catalysis is unknown. Here, we present 3 high-resolution cryo-electron microscopy structures of active Saccharomyces cerevisiae RNA Pol II elongation complexes in distinct catalytic states: two pre-catalysis states at 1.96 Å and 2.26 Å resolution and a post-catalysis state at 2.33 Å resolution. Each structure contains over 700–1,350 ordered water molecules, many located at functionally critical positions. Comparative analysis shows that these waters play essential roles in proton-transfer steps during RNA Pol II catalysis, facilitating substrate recognition and trigger-loop folding during nucleotide addition. Strikingly, these waters are conserved between prokaryotic and eukaryotic transcription machineries (see Mueller and Darst). These findings provide unprecedented mechanistic insights into RNA Pol II catalysis and reveal vital and evolutionarily conserved roles of water molecules in transcription.
|
Apr 2026
|
|
Krios IV-Titan Krios IV at Diamond
|
Rosie M.
Mundy
,
Kasim
Waraich
,
Emily A.
Bates
,
Pierre J.
Rizkallah
,
Alexander T.
Baker
,
Mark T.
Young
,
Edward
Morris
,
Paula C. A.
Da Fonseca
,
Carly M.
Bliss
,
David
Matthews
,
David
Bhella
,
Alan L.
Parker
Diamond Proposal Number(s):
[31827]
Open Access
Abstract: Adenoviruses are widely used as vectors for subunit vaccines and oncolytic therapies. Efficient vectors must infect target cells and deliver therapeutic transgenes at high levels. Species D adenoviruses, such as human adenovirus type 10 (HAdV-D10), are promising candidates due to low seroprevalence in humans. Here, we present the cryo-electron microscopy structure of the HAdV-D10 capsid alongside transcriptomic profiling of infected cells to inform vector development. The fiber shaft, essential for cell entry, was resolved at 10 Å, revealing a previously uncharacterized ‘umbrella’ motif. Viral transcript analysis using an ORF-centric pipeline uncovered key differences from HAdV-C5, including abundant expression of a transcript encoding a protein equivalent to mature protein VII. These findings highlight the importance of detailed vector characterization prior to clinical translation and support the advancement of HAdV-D10 as a next-generation platform for gene delivery and vaccine development.
|
Apr 2026
|
|
Krios IV-Titan Krios IV at Diamond
|
Diamond Proposal Number(s):
[34312]
Open Access
Abstract: Microtubule nucleation by the γ-tubulin ring complex (γTuRC) is spatiotemporally regulated and in higher eukaryotes is thought to involve a transition from an inactive open to an active closed conformation that matches the microtubule geometry. However, γTuRC activators only promote a partially closed conformation, raising the question of whether complete closure is required for activation. Combining in vitro nucleation assays and cryo-EM, we find that centrosomin motif 1 (CM1), a conserved element of several γTuRC regulators, potently accelerates human γTuRC-mediated microtubule nucleation by facilitating complete closure of γTuRC as the nascent microtubule assembles. A 3.7 Å cryo-EM structure identifies the γTuRC latch and several interactions involved in conformational closure. Notably, the distinct subunits that keep γTuRC open and inactive in higher eukaryotes also participate in its closure and activation. This work provides additional insight into the logic of the human γTuRC architecture and its activation by CM1.
|
Mar 2026
|
|
Krios II-Titan Krios II at Diamond
Krios III-Titan Krios III at Diamond
Krios IV-Titan Krios IV at Diamond
|
Diamond Proposal Number(s):
[19435, 18258]
Open Access
Abstract: Type IV pili are long, filamentous structures that extend from bacterial cell surfaces, enabling cells to respond to changing environments and facilitating genome plasticity. Thermus thermophilus HB27 produces two different type IV pili, each exhibiting distinct structural and functional properties. Here, we combine cryo-electron tomography, mutagenesis, and AlphaFold predictions to generate hypothetical in situ models of the T. thermophilus type IV pilus assembly machinery. Using single-particle cryo-electron microscopy, we determine structures of both filament types, enabling modelling of their surface glycans. Molecular dynamics simulations further reveal the flexibility of these glycans on extrusion. Integration of the filament structures with our hypothetical model of the assembly machinery offers a framework for further dissecting T4P architecture and biogenesis.
|
Mar 2026
|
|
Krios III-Titan Krios III at Diamond
Krios IV-Titan Krios IV at Diamond
|
Donghwi
Ko
,
Raili
Ruonala
,
Alexandre
Faille
,
Eva
Hellmann
,
Hanna
Help
,
Huili
Liu
,
Ronni
Nielsen
,
Anders
Haakonsson
,
Nuria
De Diego
,
Anja
Paatero
,
Mariia V.
Shcherbii
,
Karolina
Stefanowicz
,
Sanja
Ćavar Zeljković
,
Tine
Drud Lundager Rasmussen
,
Ondrej
Novak
,
Zsuzsanna
Bodi
,
Gugan
Eswaran
,
Brecht
Wybouw
,
Matthieu
Bourdon
,
Cristina
Urbez
,
Xiaonan
Liu
,
Kari
Salokas
,
Tiina
Öhman
,
Tanya
Waldie
,
Petri
Törönen
,
Sedeer
El-Showk
,
Martin
Balcerowicz
,
Fabrice
Besnard
,
Xiaomin
Liu
,
Patrick
Perkins
,
Serina
Mazzoni-Putman
,
Julia P.
Vainonen
,
Maija
Sierla
,
Mikko J.
Frilander
,
Susanne
Mandrup
,
Teva
Vernoux
,
Karin
Ljung
,
Alejandro
Ferrando
,
Miguel A.
Blazquez
,
Liisa
Holm
,
Rupert
Fray
,
Markku
Varjosalo
,
Ottoline
Leyser
,
Ville O.
Paavilainen
,
Ari Pekka
Mähönen
,
Anna
Stepanova
,
Jose
Alonso
,
Steffen
Heber
,
Robert
Malinowski
,
Finn
Kirpekar
,
Alan J.
Warren
,
Ykä
Helariutta
Diamond Proposal Number(s):
[37678]
Abstract: Polyamines are often associated with ribosomes and are thought to stabilize their integrity. In Arabidopsis, the polyamine thermospermine (tSpm) affects xylem cell fate. tSpm induces translation of SUPPRESSOR-OF-ACAULIS51 (SAC51) and SAC51-LIKEs (SACLs), which inhibit heterodimerization of the xylem development proteins LONESOME-HIGHWAY (LHW) and TARGET-OF-MONOPTEROS5. Here, we report a methyltransferase, OVERACHIEVER, that methylates the peptidyl transferase center of the 25S ribosomal RNA (rRNA). Residue m3U2952 promotes functional tSpm binding to a specific site connecting the P-site transfer RNA (tRNA) with rRNA residues in the peptidyl transferase center. This interaction enhances the translation of SACLs but inhibits that of LHW. Our study uncovers the dependency between a conserved rRNA base methylation and a polyamine in orchestrating cell fate decisions, highlighting a role for the ribosome chemical landscape in translational regulation.
|
Feb 2026
|
|
Krios I-Titan Krios I at Diamond
Krios IV-Titan Krios IV at Diamond
|
Mingda
Ye
,
Zhu
Liang
,
Daming
Zhou
,
Ashley C. W.
Pike
,
Siyi
Wang
,
Dong
Wang
,
Souvika
Bakshi
,
Laurent
Brooke
,
Eleanor P.
Williams
,
Jonathan M.
Elkins
,
Benedikt M.
Kessler
,
David I.
Stuart
,
David B.
Sauer
Diamond Proposal Number(s):
[28713]
Open Access
Abstract: Arginine, lysine, and ornithine are critical to several fundamental aspects of organismal physiology, including protein structure and function, the urea cycle, and intracellular signaling. These cationic amino acids are imported by several membrane transporters, most notably the Cationic Amino acid Transporters (CATs) in the SLC7 family. Of these, CAT1 is also the receptor for two orthoretroviruses, and determines the host tropism for these viruses. Here, using a combination of CryoEM and in vitro biochemical techniques, we characterize the substrate recognition and transport of CAT1 from Mus musculus. Further, by determining the structures of MmCAT1 in complex with the receptor binding domain from the Friend Murine Leukemia Virus, we identify the key structural interactions that determine the virus’ rodent-specific tropism.
|
Feb 2026
|
|
Krios IV-Titan Krios IV at Diamond
|
Diamond Proposal Number(s):
[36186]
Abstract: Ordered protein-based assemblies are increasingly desirable for materials science, but the design of new materials remains challenging and requires considerable effort. Crystal lattice contact modulation enables rapid rational design of an assortment of structurally diverse constructs with crystalline order. Targeted disruption of crystal contacts and directional growth pre-biasing allows for restricting crystal lattice growth in selected directions, resulting in lower-dimension assemblies with parent crystal structural features. Two-dimensional crystals, one-dimensional fibres, flexible ribbons, and single-walled nanotubes based on tetratricopeptide repeat proteins were constructed from reengineered 3D crystal lattices. The large library of available crystal structures provides an abundance of engineering targets, promising to make crystal contact engineering a rapid and attractive approach for the design of ordered supramolecular protein assemblies.
|
Oct 2025
|
|
Krios IV-Titan Krios IV at Diamond
|
Diamond Proposal Number(s):
[34172]
Open Access
Abstract: Plasmodium falciparum is a eukaryotic pathogen responsible for the majority of malaria-related fatalities. Plasmodium belongs to the phylum Apicomplexa and, like most members of this phylum, contains a non-photosynthetic plastid called the apicoplast. The apicoplast has its own genome, replicated by a dedicated replisome. Unlike other cellular replisomes, the apicoplast replisome uses a single DNA polymerase (apPol). This suggests that apPol can multitask and catalyse both replicative and lesion bypass synthesis. Replicative synthesis relies on a restrictive active site for high accuracy while lesion bypass typically requires an open active site. This raises the question: how does apPol combine the structural features of multiple DNA polymerases in a single protein? Using single-particle electron cryomicroscopy (cryoEM), we have solved the structures of apPol bound to its undamaged DNA and nucleotide substrates in five pre-chemistry conformational states. We found that apPol can accommodate a nascent base pair with the fingers in an open configuration, which might facilitate the lesion bypass activity. In the fingers-open state, we identified a nascent base pair checkpoint that preferentially selects Watson–Crick base pairs, an essential requirement for replicative synthesis. Taken together, these structural features might explain how apPol balances replicative and lesion bypass synthesis.
|
Oct 2025
|
|
