Manipulation of Host Cholesterol by SARS-CoV-2.

SARS-CoV-2 infection is associated with alterations in host lipid metabolism, including disruptions in cholesterol homeostasis. However, the specific mechanisms by which viral proteins influence cholesterol remain incompletely understood. Here, we report that SARS-CoV-2 infection induces cholesterol sequestration within lysosomes, with the viral protein ORF3a identified as the primary driver of this effect.

Calcium signaling from damaged lysosomes induces cytoprotective stress granules.

Lysosomal damage induces stress granule (SG) formation. However, the importance of SGs in determining cell fate and the precise mechanisms that mediate SG formation in response to lysosomal damage remain unclear. Here, we describe a novel calcium-dependent pathway controlling SG formation, which promotes cell survival during lysosomal damage.

Torsional Twist of the SARS-CoV and SARS-CoV-2 SUD-N and SUD-M domains.

Coronavirus non-structural protein 3 (nsp3) forms hexameric crowns of pores in the double membrane vacuole that houses the replication-transcription complex. Nsp3 in SARS-like viruses has three unique domains absent in other coronavirus nsp3 proteins. Two of these, SUD-N (Macrodomain 2) and SUD-M (Macrodomain 3), form two lobes connected by a peptide linker and an interdomain disulfide bridge.

MARTX toxin processing and degradation of cellular Rab GTPases by the cytotoxic effector Makes Caterpillars Floppy.

causes life-threatening wound and gastrointestinal infections, mediated primarily by the production of a Multifunctional-Autoprocessing Repeats-In-Toxin (MARTX) toxin. The most commonly present MARTX effector domain, the Makes Caterpillars Floppy-like (MCF) toxin, is a cysteine protease stimulated by host adenosine diphosphate (ADP) ribosylation factors (ARFs) to autoprocess. Here, we show processed MCF then binds and cleaves host s-related proteins in rain (Rab) guanosine triphosphatases within their C-terminal tails resulting in Rab degradation.

A mechanism that transduces lysosomal damage signals to stress granule formation for cell survival.

Lysosomal damage poses a significant threat to cell survival. Our previous work has reported that lysosomal damage induces stress granule (SG) formation. However, the importance of SG formation in determining cell fate and the precise mechanisms through which lysosomal damage triggers SG formation remains unclear.

A high-throughput structural system biology approach to increase structure representation of proteins from .

causes life-threatening gastrointestinal infections. It is a high-risk pathogen due to a lack of effective treatments, antimicrobial resistance, and a poorly conserved genomic core. Herein, we report 30 X-ray structures from a structure genomics pipeline spanning 13 years, representing 10.

Discovery of a Druggable, Cryptic Pocket in SARS-CoV-2 nsp16 Using Allosteric Inhibitors.

A collaborative, open-science team undertook discovery of novel small molecule inhibitors of the SARS-CoV-2 nsp16-nsp10 2'--methyltransferase using a high throughput screening approach with the potential to reveal new inhibition strategies. This screen yielded compound , a ligand possessing an electron-deficient double bond, as an inhibitor of SARS-CoV-2 nsp16 activity. Surprisingly, X-ray crystal structures revealed that covalently binds within a previously unrecognized cryptic pocket near the -adenosylmethionine binding cleft in a manner that prevents occupation by -adenosylmethionine.

Serodominant SARS-CoV-2 Nucleocapsid Peptides Map to Unstructured Protein Regions.

The SARS-CoV-2 nucleocapsid (N) protein is highly immunogenic, and anti-N antibodies are commonly used as markers for prior infection. While several studies have examined or predicted the antigenic regions of N, these have lacked consensus and structural context. Using COVID-19 patient sera to probe an overlapping peptide array, we identified six public and four private epitope regions across N, some of which are unique to this study.

MARTX toxin processing and degradation of cellular Rab GTPases by the cytotoxic effector Makes Caterpillars Floppy.

causes life threatening infections dependent upon the effectors released from the Multifunctional-Autoprocessing Repeats-In-Toxin (MARTX) toxin. The Makes Caterpillars Floppy-like (MCF) cysteine protease effector is activated by host ADP ribosylation factors (ARFs), although the targets of processing activity were unknown. In this study we show MCF binds Ras-related proteins in brain (Rab) GTPases at the same interface occupied by ARFs and then cleaves and/or degrades 24 distinct members of the Rab GTPases family.

Functional and Structural Characterization of OXA-935, a Novel OXA-10-Family β-Lactamase from Pseudomonas aeruginosa.

Resistance to antipseudomonal penicillins and cephalosporins is often driven by the overproduction of the intrinsic β-lactamase AmpC. However, OXA-10-family β-lactamases are a rich source of resistance in Pseudomonas aeruginosa. OXA β-lactamases have a propensity for mutation that leads to extended spectrum cephalosporinase and carbapenemase activity.

Mn coordinates Cap-0-RNA to align substrates for efficient 2'--methyl transfer by SARS-CoV-2 nsp16.

Capping of viral messenger RNAs is essential for efficient translation, for virus replication, and for preventing detection by the host cell innate response system. The SARS-CoV-2 genome encodes the 2'--methyltransferase nsp16, which, when bound to the coactivator nsp10, uses -adenosylmethionine (SAM) as a donor to transfer a methyl group to the first ribonucleotide of the mRNA in the final step of viral mRNA capping. Here, we provide biochemical and structural evidence that this reaction requires divalent cations, preferably Mn, and a coronavirus-specific four-residue insert.

2'-O methylation of RNA cap in SARS-CoV-2 captured by serial crystallography.

The genome of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) coronavirus has a capping modification at the 5'-untranslated region (UTR) to prevent its degradation by host nucleases. These modifications are performed by the Nsp10/14 and Nsp10/16 heterodimers using S-adenosylmethionine as the methyl donor. Nsp10/16 heterodimer is responsible for the methylation at the ribose 2'-O position of the first nucleotide.

Age-Dependent Decline in Synaptic Mitochondrial Function Is Exacerbated in Vulnerable Brain Regions of Female 3xTg-AD Mice.

Synaptic aging has been associated with neuronal circuit dysfunction and cognitive decline. Reduced mitochondrial function may be an early event that compromises synaptic integrity and neurotransmission in vulnerable brain regions during physiological and pathological aging. Thus, we aimed to measure mitochondrial function in synapses from three brain regions at two different ages in the 3xTg-AD mouse model and in wild mice.

High-resolution structures of the SARS-CoV-2 2'--methyltransferase reveal strategies for structure-based inhibitor design.

There are currently no antiviral therapies specific for SARS-CoV-2, the virus responsible for the global pandemic disease COVID-19. To facilitate structure-based drug design, we conducted an x-ray crystallographic study of the SARS-CoV-2 nsp16-nsp10 2'--methyltransferase complex, which methylates Cap-0 viral mRNAs to improve viral protein translation and to avoid host immune detection. We determined the structures for nsp16-nsp10 heterodimers bound to the methyl donor -adenosylmethionine (SAM), the reaction product -adenosylhomocysteine (SAH), or the SAH analog sinefungin (SFG).

The crystal structure of nsp10-nsp16 heterodimer from SARS-CoV-2 in complex with S-adenosylmethionine.

SARS-CoV-2 is a member of the coronaviridae family and is the etiological agent of the respiratory Coronavirus Disease 2019. The virus has spread rapidly around the world resulting in over two million cases and nearly 150,000 deaths as of April 17, 2020. Since no treatments or vaccines are available to treat COVID-19 and SARS-CoV-2, respiratory complications derived from the infections have overwhelmed healthcare systems around the world.

The aerobic respiratory chain of Pseudomonas aeruginosa cultured in artificial urine media: Role of NQR and terminal oxidases.

Pseudomonas aeruginosa is a Gram-negative γ-proteobacterium that forms part of the normal human microbiota and it is also an opportunistic pathogen, responsible for 30% of all nosocomial urinary tract infections. P. aeruginosa carries a highly branched respiratory chain that allows the colonization of many environments, such as the urinary tract, catheters and other medical devices.

Comparison of metal-bound and unbound structures of aminopeptidase B proteins from Escherichia coli and Yersinia pestis.

Protein degradation by aminopeptidases is involved in bacterial responses to stress. Escherichia coli produces two metal-dependent M17 family leucine aminopeptidases (LAPs), aminopeptidase A (PepA) and aminopeptidase B (PepB). Several structures have been solved for PepA as well as other bacterial M17 peptidases.

Structure of galactarate dehydratase, a new fold in an enolase involved in bacterial fitness after antibiotic treatment.

Galactarate dehydratase (GarD) is the first enzyme in the galactarate/glucarate pathway and catalyzes the dehydration of galactarate to 3-keto-5-dehydroxygalactarate. This protein is known to increase colonization fitness of intestinal pathogens in antibiotic-treated mice and to promote bacterial survival during stress. The galactarate/glucarate pathway is widespread in bacteria, but not in humans, and thus could be a target to develop new inhibitors for use in combination therapy to combat antibiotic resistance.

Structural comparison of p-hydroxybenzoate hydroxylase (PobA) from Pseudomonas putida with PobA from other Pseudomonas spp. and other monooxygenases.

The crystal structure is reported of p-hydroxybenzoate hydroxylase (PobA) from Pseudomonas putida, a possible drug target to combat tetracycline resistance, in complex with flavin adenine dinucleotide (FAD). The structure was refined at 2.2 Å resolution with four polypeptide chains in the asymmetric unit.

Cholesterol burden in the liver induces mitochondrial dynamic changes and resistance to apoptosis.

Non-alcoholic fatty liver disease (NAFLD) encompasses a broad spectrum of histopathological changes ranging from non-inflammatory intracellular fat deposition to non-alcoholic steatohepatitis (NASH), which may progress into hepatic fibrosis, cirrhosis, or hepatocellular carcinoma. Recent data suggest that impaired hepatic cholesterol homeostasis and its accumulation are relevant to the pathogenesis of NAFLD/NASH. Despite a vital physiological function of cholesterol, mitochondrial dysfunction is an important consequence of dietary-induced hypercholesterolemia and was, subsequently, linked to many pathophysiological conditions.

Characterization of the NQR complex, a bacterial proton pump with roles in autopoisoning resistance.

is a Gram-negative bacterium responsible for a large number of nosocomial infections. The respiratory chain contains the ion-pumping NADH:ubiquinone oxidoreductase (NQR). This enzyme couples the transfer of electrons from NADH to ubiquinone to the pumping of sodium ions across the cell membrane, generating a gradient that drives essential cellular processes in many bacteria.

The role of glycolysis-derived hexose phosphates in the induction of the Crabtree effect.

Evidence for the Crabtree effect was first reported by H. Crabtree in 1929 and is defined as the glucose-induced decrease of cellular respiratory flux. This effect was observed in tumor cells and was not detected in most non-tumor cells.

Alternative mitochondrial respiratory chains from two crustaceans: Artemia franciscana nauplii and the white shrimp, Litopenaeus vannamei.

Mitochondrial ATP is synthesized by coupling between the electron transport chain and complex V. In contrast, physiological uncoupling of these processes allows mitochondria to consume oxygen at high rates without ATP synthesis. Such uncoupling mechanisms prevent reactive oxygen species overproduction.

Dynamic energy dependency of on host cell metabolism during intracellular growth: Role of sodium-based energetics in chlamydial ATP generation.

is an obligate intracellular human pathogen responsible for the most prevalent sexually-transmitted infection in the world. For decades has been considered an "energy parasite" that relies entirely on the uptake of ATP from the host cell. The genomic data suggest that respiratory chain could produce a sodium gradient that may sustain the energetic demands required for its rapid multiplication.