scholarly journals Chemoproteomics of microbiota metabolites reveals small-molecule agonists for orphan receptor GPRC5A

2021 ◽  
Author(s):  
Xiaohui Zhao ◽  
Kathryn R. Stein ◽  
Victor Chen ◽  
Matthew E. Griffin ◽  
Howard C. Hang
Keyword(s):  
Small Molecule ◽  
Biochemical Characterization ◽  
Bacterial Species ◽  
Host Cells ◽  
Orphan Receptor ◽  
Amino Acid Residues ◽  
Protein Targets ◽  
Functional Studies ◽  
Functional Profiling ◽  
G Protein Coupled

The microbiota generates diverse metabolites that can engage multiple pathways to modulate host physiology and disease, but their protein targets and mechanism(s) of action have not been fully elucidated. To address this challenge, we focused on indole-3-acetic acid (IAA), a prominent microbiota metabolite, and developed IAA-based chemical reporters for proteomic studies. We discovered that IAA interacts with many proteins in host cells, including small-molecule transporters, receptors and metabolic enzymes. Notably, our functional studies revealed that IAA binds to orphan G protein-coupled receptors such as GPRC5A, but only aromatic monoamines were capable of inducing GPRC5A signaling. Functional profiling of microbiota uncovered specific bacterial species and enzymes that generate GPRC5A agonists. Finally, biochemical characterization of GPRC5A activation identified more potent synthetic agonists as well as key amino acid residues involved in ligand binding. These studies highlight the utility of chemoproteomics to dissect protein targets and mechanisms of action for microbiota metabolites.

scholarly journals Dengue and Zika virus infection are impaired by small molecule ER proteostasis regulator 147 in an ATF6-independent manner

2020 ◽  
Author(s):  
Katherine M. Almasy ◽  
Jonathan P. Davies ◽  
Samantha M. Lisy ◽  
Reyhaneh Tirgar ◽  
Sirena C. Tran ◽  
...  
Keyword(s):  
Small Molecule ◽  
Zika Virus ◽  
Antiviral Agent ◽  
Host Cells ◽  
Human Pathogens ◽  
Independent Manner ◽  
Protein Targets ◽  
Disulfide Isomerases ◽  
Multiple Strains ◽  
The Unfolded Protein Response

ABSTRACTFlaviviruses, including Dengue and Zika, are widespread human pathogens, however, no broadly active therapeutics exist to fight infection. Here, we establish the recently discovered pharmacologic modulator of ER proteostasis 147 as an effective host-centered antiviral strategy. Compound 147 reduces infection by attenuating viral replication without causing toxicity in host cells. 147 is a preferential activator of the ATF6 pathway of the unfolded protein response, which requires targeting of cysteine residues primarily on protein disulfide isomerases (PDIs). We find that the antiviral activity of 147 is independent of ATF6 induction but does require modification of reactive thiols on protein targets. Targeting PDIs using RNAi and other PDI small molecule inhibitors was unable to recapitulate the antiviral effects, suggesting additional identified protein targets of 147 may mediate the activity. Importantly, 147 can impair infection of multiple strains of Dengue and Zika virus, indicating that it is suitable as a broad-spectrum antiviral agent.

Strategies to identify ligands for orphan G-protein-coupled receptors

2002 ◽  
Vol 30 (4) ◽  
pp. 789-793 ◽  
Author(s):  
G. Milligan
Keyword(s):  
Drug Design ◽  
G Protein ◽  
Small Molecule ◽  
Drug Targets ◽  
G Protein Coupled Receptors ◽  
Orphan Receptors ◽  
Protein Targets ◽  
Small Molecule Drug ◽  
Natural Ligands ◽  
G Protein Coupled

G-protein-coupled receptors are the most tractable class of protein targets for small molecule drug design. Sequencing of the human genome allied to bio-informatic analysis has identified a large number of putative receptors for which the natural ligands remain undefined. A range of currently employed and developing strategies to identify ligands that interact with these orphan receptors and to validate them as drug targets are described and discussed.

Dicarbonyl derived post-translational modifications: chemistry bridging biology and aging-related disease

2020 ◽  
Vol 64 (1) ◽  
pp. 97-110
Author(s):  
Christian Sibbersen ◽  
Mogens Johannsen
Keyword(s):  
Mass Spectrometry ◽  
Future Research ◽  
Amino Acid Residues ◽  
Post Translational Modification ◽  
Dicarbonyl Compounds ◽  
Protein Targets ◽  
Post Translational Modifications ◽  
Reactive Protein ◽  
Glycation End Products ◽  
Direct Mass Spectrometry

Abstract In living systems, nucleophilic amino acid residues are prone to non-enzymatic post-translational modification by electrophiles. α-Dicarbonyl compounds are a special type of electrophiles that can react irreversibly with lysine, arginine, and cysteine residues via complex mechanisms to form post-translational modifications known as advanced glycation end-products (AGEs). Glyoxal, methylglyoxal, and 3-deoxyglucosone are the major endogenous dicarbonyls, with methylglyoxal being the most well-studied. There are several routes that lead to the formation of dicarbonyl compounds, most originating from glucose and glucose metabolism, such as the non-enzymatic decomposition of glycolytic intermediates and fructosyl amines. Although dicarbonyls are removed continuously mainly via the glyoxalase system, several conditions lead to an increase in dicarbonyl concentration and thereby AGE formation. AGEs have been implicated in diabetes and aging-related diseases, and for this reason the elucidation of their structure as well as protein targets is of great interest. Though the dicarbonyls and reactive protein side chains are of relatively simple nature, the structures of the adducts as well as their mechanism of formation are not that trivial. Furthermore, detection of sites of modification can be demanding and current best practices rely on either direct mass spectrometry or various methods of enrichment based on antibodies or click chemistry followed by mass spectrometry. Future research into the structure of these adducts and protein targets of dicarbonyl compounds may improve the understanding of how the mechanisms of diabetes and aging-related physiological damage occur.

scholarly journals Cutting Edge: Identification of the Orphan Receptor G-Protein-Coupled Receptor 2 as CCR10, a Specific Receptor for the Chemokine ESkine

2000 ◽  
Vol 164 (7) ◽  
pp. 3460-3464 ◽  
Author(s):  
David I. Jarmin ◽  
Miriam Rits ◽  
Dalena Bota ◽  
Norma P. Gerard ◽  
Gerard J. Graham ◽  
...  
Keyword(s):  
G Protein ◽  
Cutting Edge ◽  
Orphan Receptor ◽  
Specific Receptor ◽  
G Protein Coupled Receptor ◽  
G Protein Coupled

scholarly journals Monoassociation with bacterial isolates reveals the role of colonization, community complexity and abundance on locomotor behavior in larval zebrafish

2021 ◽  
Vol 3 (1) ◽  
Author(s):  
Chelsea A. Weitekamp ◽  
Allison Kvasnicka ◽  
Scott P. Keely ◽  
Nichole E. Brinkman ◽  
Xia Meng Howey ◽  
...  
Keyword(s):  
Total Concentration ◽  
Bacterial Species ◽  
Locomotor Behavior ◽  
Host Cells ◽  
Individual Species ◽  
Zebrafish Model ◽  
Associated Bacteria ◽  
Larval Zebrafish ◽  
Toxicological Studies ◽  
Community Complexity

Abstract Background Across taxa, animals with depleted intestinal microbiomes show disrupted behavioral phenotypes. Axenic (i.e., microbe-free) mice, zebrafish, and fruit flies exhibit increased locomotor behavior, or hyperactivity. The mechanism through which bacteria interact with host cells to trigger normal neurobehavioral development in larval zebrafish is not well understood. Here, we monoassociated zebrafish with either one of six different zebrafish-associated bacteria, mixtures of these host-associates, or with an environmental bacterial isolate. Results As predicted, the axenic cohort was hyperactive. Monoassociation with three different host-associated bacterial species, as well as with the mixtures, resulted in control-like locomotor behavior. Monoassociation with one host-associate and the environmental isolate resulted in the hyperactive phenotype characteristic of axenic larvae, while monoassociation with two other host-associated bacteria partially blocked this phenotype. Furthermore, we found an inverse relationship between the total concentration of bacteria per larvae and locomotor behavior. Lastly, in the axenic and associated cohorts, but not in the larvae with complex communities, we detected unexpected bacteria, some of which may be present as facultative predators. Conclusions These data support a growing body of evidence that individual species of bacteria can have different effects on host behavior, potentially related to their success at intestinal colonization. Specific to the zebrafish model, our results suggest that differences in the composition of microbes in fish facilities could affect the results of behavioral assays within pharmacological and toxicological studies.

scholarly journals T6SS Mediated Stress Responses for Bacterial Environmental Survival and Host Adaptation

2021 ◽  
Vol 22 (2) ◽  
pp. 478
Author(s):  
Kai-Wei Yu ◽  
Peng Xue ◽  
Yang Fu ◽  
Liang Yang
Keyword(s):  
Protein Secretion ◽  
Stress Responses ◽  
Current Knowledge ◽  
Bacterial Species ◽  
Host Cells ◽  
Interspecies Interactions ◽  
Type Vi Secretion System ◽  
Membrane Complex ◽  
Type Vi Secretion ◽  
Bacterial Type

The bacterial type VI secretion system (T6SS) is a protein secretion apparatus widely distributed in Gram-negative bacterial species. Many bacterial pathogens employ T6SS to compete with the host and to coordinate the invasion process. The T6SS apparatus consists of a membrane complex and an inner tail tube-like structure that is surrounded by a contractile sheath and capped with a spike complex. A series of antibacterial or antieukaryotic effectors is delivered by the puncturing device consisting of a Hcp tube decorated by the VgrG/PAAR complex into the target following the contraction of the TssB/C sheath, which often leads to damage and death of the competitor and/or host cells. As a tool for protein secretion and interspecies interactions, T6SS can be triggered by many different mechanisms to respond to various physiological conditions. This review summarizes our current knowledge of T6SS in coordinating bacterial stress responses against the unfavorable environmental and host conditions.

scholarly journals Legionella pneumophila DotU and IcmF Are Required for Stability of the Dot/Icm Complex

2004 ◽  
Vol 72 (10) ◽  
pp. 5983-5992 ◽  
Author(s):  
Jessica A. Sexton ◽  
Jennifer L. Miller ◽  
Aki Yoneda ◽  
Thomas E. Kehl-Fie ◽  
Joseph P. Vogel
Keyword(s):  
Cell Surface ◽  
Legionella Pneumophila ◽  
Bacterial Species ◽  
Wide Distribution ◽  
Host Cells ◽  
Growth Defect ◽  
Intracellular Growth ◽  
Homologous Proteins ◽  
Type Iv ◽  
Cell Surface Structure

ABSTRACT Legionella pneumophila utilizes a type IV secretion system (T4SS) encoded by 26 dot/icm genes to replicate inside host cells and cause disease. In contrast to all other L. pneumophila dot/icm genes, dotU and icmF have homologs in a wide variety of gram-negative bacteria, none of which possess a T4SS. Instead, dotU and icmF orthologs are linked to a locus encoding a conserved cluster of proteins designated IcmF-associated homologous proteins, which has been proposed to constitute a novel cell surface structure. We show here that dotU is partially required for L. pneumophila intracellular growth, similar to the known requirement for icmF. In addition, we show that dotU and icmF are necessary for optimal plasmid transfer and sodium sensitivity, two additional phenotypes associated with a functional Dot/Icm complex. We found that these effects are due to the destabilization of the T4SS at the transition into the stationary phase, the point at which L. pneumophila becomes virulent. Specifically, three Dot proteins (DotH, DotG, and DotF) exhibit decreased stability in a ΔdotU ΔicmF strain. Furthermore, overexpression of just one of these proteins, DotH, is sufficient to suppress the intracellular growth defect of the ΔdotU ΔicmF mutant. This suggests a model where the DotU and IcmF proteins serve to prevent DotH degradation and therefore function to stabilize the L. pneumophila T4SS. Due to their wide distribution among bacterial species and their genetic linkage to known or predicted cell surface structures, we propose that this function in complex stabilization may be broadly conserved.

scholarly journals Characterization of the G protein-coupled receptor family SREB across fish evolution

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Timothy S. Breton ◽  
William G. B. Sampson ◽  
Benjamin Clifford ◽  
Anyssa M. Phaneuf ◽  
Ilze Smidt ◽  
...  
Keyword(s):  
G Protein ◽  
Genomic Structure ◽  
Integrated Approach ◽  
Orphan Receptor ◽  
Specific Gene ◽  
Testicular Development ◽  
Brain Expression ◽  
And Function ◽  
G Protein Coupled ◽  
Receptor Family

AbstractThe SREB (Super-conserved Receptors Expressed in Brain) family of G protein-coupled receptors is highly conserved across vertebrates and consists of three members: SREB1 (orphan receptor GPR27), SREB2 (GPR85), and SREB3 (GPR173). Ligands for these receptors are largely unknown or only recently identified, and functions for all three are still beginning to be understood, including roles in glucose homeostasis, neurogenesis, and hypothalamic control of reproduction. In addition to the brain, all three are expressed in gonads, but relatively few studies have focused on this, especially in non-mammalian models or in an integrated approach across the entire receptor family. The purpose of this study was to more fully characterize sreb genes in fish, using comparative genomics and gonadal expression analyses in five diverse ray-finned (Actinopterygii) species across evolution. Several unique characteristics were identified in fish, including: (1) a novel, fourth euteleost-specific gene (sreb3b or gpr173b) that likely emerged from a copy of sreb3 in a separate event after the teleost whole genome duplication, (2) sreb3a gene loss in Order Cyprinodontiformes, and (3) expression differences between a gar species and teleosts. Overall, gonadal patterns suggested an important role for all sreb genes in teleost testicular development, while gar were characterized by greater ovarian expression that may reflect similar roles to mammals. The novel sreb3b gene was also characterized by several unique features, including divergent but highly conserved amino acid positions, and elevated brain expression in puffer (Dichotomyctere nigroviridis) that more closely matched sreb2, not sreb3a. These results demonstrate that SREBs may differ among vertebrates in genomic structure and function, and more research is needed to better understand these roles in fish.

scholarly journals A small molecule compound with an indole moiety inhibits the main protease of SARS-CoV-2 and blocks virus replication

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Shin-ichiro Hattori ◽  
Nobuyo Higashi-Kuwata ◽  
Hironori Hayashi ◽  
Srinivasa Rao Allu ◽  
Jakka Raghavaiah ◽  
...  
Keyword(s):  
Small Molecule ◽  
Covalent Bond ◽  
Amino Acid Residues ◽  
X Ray ◽  
Viral Breakthrough ◽  
Main Protease ◽  
Small Molecule Compound ◽  
Polar Interactions ◽  
High Concentrations

AbstractExcept remdesivir, no specific antivirals for SARS-CoV-2 infection are currently available. Here, we characterize two small-molecule-compounds, named GRL-1720 and 5h, containing an indoline and indole moiety, respectively, which target the SARS-CoV-2 main protease (Mpro). We use VeroE6 cell-based assays with RNA-qPCR, cytopathic assays, and immunocytochemistry and show both compounds to block the infectivity of SARS-CoV-2 with EC50 values of 15 ± 4 and 4.2 ± 0.7 μM for GRL-1720 and 5h, respectively. Remdesivir permitted viral breakthrough at high concentrations; however, compound 5h completely blocks SARS-CoV-2 infection in vitro without viral breakthrough or detectable cytotoxicity. Combination of 5h and remdesivir exhibits synergism against SARS-CoV-2. Additional X-ray structural analysis show that 5h forms a covalent bond with Mpro and makes polar interactions with multiple active site amino acid residues. The present data suggest that 5h might serve as a lead Mpro inhibitor for the development of therapeutics for SARS-CoV-2 infection.

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