Antibodies disrupt bacterial adhesion by ligand mimicry and allosteric interference.

A critical step in infections is the attachment of many microorganisms to host cells using lectins that bind surface glycans, making lectins promising antimicrobial targets. Upon binding mannosylated glycans, FimH, the most studied lectin adhesin of type 1 fimbriae in , undergoes an allosteric transition from an inactive to an active conformation that can act as a catch-bond. Monoclonal antibodies that alter FimH glycan binding in various ways are available, but the mechanisms of these antibodies remain unclear.

Isolation and escape mapping of broadly neutralizing antibodies against emerging delta-coronaviruses.

Porcine delta-coronavirus (PDCoV) spillovers were recently detected in febrile children, underscoring the recurrent zoonoses of divergent CoVs. To date, no vaccines or specific therapeutics are approved for use in humans against PDCoV. To prepare for possible future PDCoV epidemics, we isolated PDCoV spike (S)-directed monoclonal antibodies (mAbs) from humanized mice and found that two, designated PD33 and PD41, broadly neutralized a panel of PDCoV variants.

Broadly neutralizing antibodies against emerging delta-coronaviruses.

Porcine deltacoronavirus (PDCoV) spillovers were recently detected in children with acute undifferentiated febrile illness, underscoring recurrent zoonoses of divergent coronaviruses. To date, no vaccines or specific therapeutics are approved for use in humans against PDCoV. To prepare for possible future PDCoV epidemics, we isolated human spike (S)-directed monoclonal antibodies from transgenic mice and found that two of them, designated PD33 and PD41, broadly neutralized a panel of PDCoV variants.

Light-sensitive phosphorylation regulates retinal IMPDH1 activity and filament assembly.

Inosine monophosphate dehydrogenase (IMPDH) is the rate-limiting enzyme in guanosine triphosphate (GTP) synthesis and assembles into filaments in cells, which desensitizes the enzyme to feedback inhibition and boosts nucleotide production. The vertebrate retina expresses two splice variants IMPDH1(546) and IMPDH1(595). In bovine retinas, residue S477 is preferentially phosphorylated in the dark, but the effects on IMPDH1 activity and regulation are unclear.

Broad receptor tropism and immunogenicity of a clade 3 sarbecovirus.

Although Rhinolophus bats harbor diverse clade 3 sarbecoviruses, the structural determinants of receptor tropism along with the antigenicity of their spike (S) glycoproteins remain uncharacterized. Here, we show that the African Rhinolophus bat clade 3 sarbecovirus PRD-0038 S has a broad angiotensin-converting enzyme 2 (ACE2) usage and that receptor-binding domain (RBD) mutations further expand receptor promiscuity and enable human ACE2 utilization. We determine a cryo-EM structure of the PRD-0038 RBD bound to Rhinolophus alcyone ACE2, explaining receptor tropism and highlighting differences with SARS-CoV-1 and SARS-CoV-2.

Light-sensitive phosphorylation regulates enzyme activity and filament assembly of human IMPDH1 retinal splice variants.

Inosine monophosphate dehydrogenase (IMPDH) is the rate-limiting enzyme in guanosine triphosphate (GTP) synthesis and is controlled by feedback inhibition and allosteric regulation. IMPDH assembles into micron-scale filaments in cells, which desensitizes the enzyme to feedback inhibition by GTP and boosts nucleotide production. The vertebrate retina expresses two tissue-specific splice variants IMPDH1(546) and IMPDH1(595).

Broad receptor tropism and immunogenicity of a clade 3 sarbecovirus.

Although bats harbor diverse clade 3 sarbecoviruses, the structural determinants of receptor tropism along with the antigenicity of their spike (S) glycoproteins remain uncharacterized. Here, we show that the African Rinolophus bat clade 3 sarbecovirus PRD-0038 S has a broad ACE2 usage and that RBD mutations further expand receptor promiscuity and enable human ACE2 utilization. We determined a cryoEM structure of the PRD-0038 RBD bound to ACE2, explaining receptor tropism and highlighting differences with SARS-CoV-1 and SARS-CoV-2.

Neutralization, effector function and immune imprinting of Omicron variants.

Currently circulating SARS-CoV-2 variants have acquired convergent mutations at hot spots in the receptor-binding domain (RBD) of the spike protein. The effects of these mutations on viral infection and transmission and the efficacy of vaccines and therapies remains poorly understood. Here we demonstrate that recently emerged BQ.

Human PRPS1 filaments stabilize allosteric sites to regulate activity.

The universally conserved enzyme phosphoribosyl pyrophosphate synthetase (PRPS) assembles filaments in evolutionarily diverse organisms. PRPS is a key regulator of nucleotide metabolism, and mutations in the human enzyme PRPS1 lead to a spectrum of diseases. Here we determine structures of human PRPS1 filaments in active and inhibited states, with fixed assembly contacts accommodating both conformations.

IMPDH1 retinal variants control filament architecture to tune allosteric regulation.

Inosine-5'-monophosphate dehydrogenase (IMPDH), a key regulatory enzyme in purine nucleotide biosynthesis, dynamically assembles filaments in response to changes in metabolic demand. Humans have two isoforms: IMPDH2 filaments reduce sensitivity to feedback inhibition, while IMPDH1 assembly remains uncharacterized. IMPDH1 plays a unique role in retinal metabolism, and point mutants cause blindness.

Cryo-EM structures of CTP synthase filaments reveal mechanism of pH-sensitive assembly during budding yeast starvation.

Many metabolic enzymes self-assemble into micron-scale filaments to organize and regulate metabolism. The appearance of these assemblies often coincides with large metabolic changes as in development, cancer, and stress. Yeast undergo cytoplasmic acidification upon starvation, triggering the assembly of many metabolic enzymes into filaments.

Structural basis for isoform-specific inhibition of human CTPS1.

Cytidine triphosphate synthase 1 (CTPS1) is necessary for an effective immune response, as revealed by severe immunodeficiency in CTPS1-deficient individuals [E. Martin ], [] [510], [288-292] ([2014]). CTPS1 expression is up-regulated in activated lymphocytes to expand CTP pools [E.

Structural basis for broad coronavirus neutralization.

Three highly pathogenic β-coronaviruses have crossed the animal-to-human species barrier in the past two decades: SARS-CoV, MERS-CoV and SARS-CoV-2. To evaluate the possibility of identifying antibodies with broad neutralizing activity, we isolated a monoclonal antibody, termed B6, that cross-reacts with eight β-coronavirus spike glycoproteins, including all five human-infecting β-coronaviruses. B6 broadly neutralizes entry of pseudotyped viruses from lineages A and C, but not from lineage B, and the latter includes SARS-CoV and SARS-CoV-2.

Coordination of phage genome degradation versus host genome protection by a bifunctional restriction-modification enzyme visualized by CryoEM.

Restriction enzymes that combine methylation and cleavage into a single assemblage and modify one DNA strand are capable of efficient adaptation toward novel targets. However, they must reliably cleave invasive DNA and methylate newly replicated unmodified host sites. One possible solution is to enforce a competition between slow methylation at a single unmodified host target, versus faster cleavage that requires multiple unmodified target sites in foreign DNA to be brought together in a reaction synapse.

Structural basis for broad coronavirus neutralization.

Three highly pathogenic β-coronaviruses crossed the animal-to-human species barrier in the past two decades: SARS-CoV, MERS-CoV and SARS-CoV-2. SARS-CoV-2 has infected more than 64 million people worldwide, claimed over 1.4 million lives and is responsible for the ongoing COVID-19 pandemic.

Unexpected Receptor Functional Mimicry Elucidates Activation of Coronavirus Fusion.

Recent outbreaks of severe acute respiratory syndrome and Middle East respiratory syndrome, along with the threat of a future coronavirus-mediated pandemic, underscore the importance of finding ways to combat these viruses. The trimeric spike transmembrane glycoprotein S mediates entry into host cells and is the major target of neutralizing antibodies. To understand the humoral immune response elicited upon natural infections with coronaviruses, we structurally characterized the SARS-CoV and MERS-CoV S glycoproteins in complex with neutralizing antibodies isolated from human survivors.

Transmission electron microscopy as an orthogonal method to characterize protein aggregates.

Aggregation of protein-based therapeutics is a challenging problem in the biopharmaceutical industry. Of particular concern are implications for product efficacy and clinical safety because of potentially increased immunogenicity of the aggregates. We used transmission electron microscopy (TEM) to characterize biophysical and morphological features of antibody aggregates formed upon controlled environmental stresses.

Maturation in action: CryoEM study of a viral capsid caught during expansion.

Bacteriophage HK97 maturation involves discrete intermediate particle forms, comparable to transitional states in protein folding, before reaching its mature form. The process starts by formation of a metastable prohead, poised for exothermic expansion triggered by DNA packaging. During maturation, the capsid subunit transitions from a strained to a canonical tertiary conformation and this has been postulated to be the driving mechanism for initiating expansion via switching hexameric capsomer architecture from skewed to 6-fold symmetric.

ATP-triggered conformational changes delineate substrate-binding and -folding mechanics of the GroEL chaperonin.

The chaperonin GroEL assists the folding of nascent or stress-denatured polypeptides by actions of binding and encapsulation. ATP binding initiates a series of conformational changes triggering the association of the cochaperonin GroES, followed by further large movements that eject the substrate polypeptide from hydrophobic binding sites into a GroES-capped, hydrophilic folding chamber. We used cryo-electron microscopy, statistical analysis, and flexible fitting to resolve a set of distinct GroEL-ATP conformations that can be ordered into a trajectory of domain rotation and elevation.

Nanoscale assembly in biological systems: from neuronal cytoskeletal proteins to curvature stabilizing lipids.

The review will describe experiments inspired by the rich variety of bundles and networks of interacting microtubules (MT), neurofilaments, and filamentous-actin in neurons where the nature of the interactions, structures, and structure-function correlations remain poorly understood. We describe how three-dimensional (3D) MT bundles and 2D MT bundles may assemble, in cell free systems in the presence of counter-ions, revealing structures not predicted by polyelectrolyte theories. Interestingly, experiments reveal that the neuronal protein tau, an abundant MT-associated-protein in axons, modulates the MT diameter providing insight for the control of geometric parameters in bio- nanotechnology.

Block Liposome and Nanotube Formation is a General Phenomenon of Two-Component Membranes Containing Multivalent Lipids.

We report a study on the formation of block liposomes (BLs) and nanotubes from membranes comprised of mixtures of membrane curvature-stabilizing multivalent cationic lipids MVL3(3+) and MVL5(5+) with neutral 1,2-dioleoyl-sn-glycero-3-phosphatidylcholine (DOPC). In conjunction with prior work on MVLBG2(16+), our experiments suggest that BL and nanotube formation is a general phenomenon in membranes containing multivalent lipids, thus enhancing the relevance of BLs for applications such as gene/drug storage and delivery or templating.

Automation in single-particle electron microscopy connecting the pieces.

Throughout the history of single-particle electron microscopy (EM), automated technologies have seen varying degrees of emphasis and development, usually depending upon the contemporary demands of the field. We are currently faced with increasingly sophisticated devices for specimen preparation, vast increases in the size of collected data sets, comprehensive algorithms for image processing, sophisticated tools for quality assessment, and an influx of interested scientists from outside the field who might lack the skills of experienced microscopists. This situation places automated techniques in high demand.

A toolbox for ab initio 3-D reconstructions in single-particle electron microscopy.

Structure determination of a novel macromolecular complex via single-particle electron microscopy depends upon overcoming the challenge of establishing a reliable 3-D reconstruction using only 2-D images. There are a variety of strategies that deal with this issue, but not all of them are readily accessible and straightforward to use. We have developed a "toolbox" of ab initio reconstruction techniques that provide several options for calculating 3-D volumes in an easily managed and tightly controlled work-flow that adheres to standard conventions and formats.

Block liposomes vesicles of charged lipids with distinctly shaped nanoscale sphere-, pear-, tube-, or rod-segments.

We describe the preparation and characterization of block liposomes, a new class of liquid (chain-melted) vesicles, from mixtures of the highly charged (+16 e) multivalent cationic lipid MVLBG2 and 1,2-dioleoyl-sn-glycero-3-phosphatidylcholine (DOPC). Block liposomes (BLs) consist of distinct spherical, tubular vesicles, and cylindrical micelles that remain connected, forming a single liposome. This is in contrast to typical liposome systems, where distinctly shaped liposomes are macroscopically separated.