scholarly journals Key computational findings reveal proton transfer as driving the functional cycle in the phosphate transporter PiPT

2021 ◽  
Vol 118 (25) ◽  
pp. e2101932118
Author(s):  
Yu Liu ◽  
Chenghan Li ◽  
Meghna Gupta ◽  
Nidhi Verma ◽  
Atul Kumar Johri ◽  
...  
Keyword(s):  
Lipid Synthesis ◽  
Phosphate Transport ◽  
Phosphate Transporter ◽  
Phosphate Transporters ◽  
Transport Pathways ◽  
Phosphate Release ◽  
Exit Tunnel ◽  
Free Energy Sampling ◽  
Dynamics Simulations ◽  
Major Facilitator

Phosphate is an indispensable metabolite in a wide variety of cells and is involved in nucleotide and lipid synthesis, signaling, and chemical energy storage. Proton-coupled phosphate transporters within the major facilitator family are crucial for phosphate uptake in plants and fungi. Similar proton-coupled phosphate transporters have been found in different protozoan parasites that cause human diseases, in breast cancer cells with elevated phosphate demand, in osteoclast-like cells during bone reabsorption, and in human intestinal Caco2BBE cells for phosphate homeostasis. However, the mechanism of proton-driven phosphate transport remains unclear. Here, we demonstrate in a eukaryotic, high-affinity phosphate transporter from Piriformospora indica (PiPT) that deprotonation of aspartate 324 (D324) triggers phosphate release. Quantum mechanics/molecular mechanics molecular dynamics simulations combined with free energy sampling have been employed here to identify the proton transport pathways from D324 upon the transition from the occluded structure to the inward open structure and phosphate release. The computational insights so gained are then corroborated by studies of D45N and D45E amino acid substitutions via mutagenesis experiments. Our findings confirm the function of the structurally predicted cytosolic proton exit tunnel and suggest insights into the role of the titratable phosphate substrate.

scholarly journals Structural and mechanistic basis for translation inhibition by macrolide and ketolide antibiotics

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Bertrand Beckert ◽  
Elodie C. Leroy ◽  
Shanmugapriya Sothiselvam ◽  
Lars V. Bock ◽  
Maxim S. Svetlov ◽  
...  
Keyword(s):  
Antibiotic Resistance Genes ◽  
Structural Basis ◽  
Sequence Motifs ◽  
Specific Sequence ◽  
Bacterial Ribosome ◽  
Translational Arrest ◽  
Exit Tunnel ◽  
Mechanistic Basis ◽  
Dynamics Simulations ◽  
Context Specific

AbstractMacrolides and ketolides comprise a family of clinically important antibiotics that inhibit protein synthesis by binding within the exit tunnel of the bacterial ribosome. While these antibiotics are known to interrupt translation at specific sequence motifs, with ketolides predominantly stalling at Arg/Lys-X-Arg/Lys motifs and macrolides displaying a broader specificity, a structural basis for their context-specific action has been lacking. Here, we present structures of ribosomes arrested during the synthesis of an Arg-Leu-Arg sequence by the macrolide erythromycin (ERY) and the ketolide telithromycin (TEL). Together with deep mutagenesis and molecular dynamics simulations, the structures reveal how ERY and TEL interplay with the Arg-Leu-Arg motif to induce translational arrest and illuminate the basis for the less stringent sequence-specific action of ERY over TEL. Because programmed stalling at the Arg/Lys-X-Arg/Lys motifs is used to activate expression of antibiotic resistance genes, our study also provides important insights for future development of improved macrolide antibiotics.

Novel Specific Inhibitors of Epithelial Phosphate Transport

Physiology ◽  
1988 ◽  
Vol 3 (4) ◽  
pp. 154-157
Author(s):  
SA Kempson
Keyword(s):  
Epithelial Cell ◽  
Cell Membranes ◽  
Phosphate Transport ◽  
Phosphate Transporter ◽  
Transport Function ◽  
Major Difficulty ◽  
Membrane Bound ◽  
Specific Label ◽  
Specific Inhibitors

The characteristics of phosphate transport across epithelial cell membranes and the influence of hormones and other factors have been studied in detail. Almost no information has emerged about the identity, structure, and biogenesis of the membrane-bound phosphate transporting proteins or how their transport function is regulated by intracellular events. One major difficulty has been the unavailability of a highly specific label for the phosphate transporter.

Inhibition of microsomal glucose-6-phosphate transport in human neutrophils results in apoptosis: a potential explanation for neutrophil dysfunction in glycogen storage disease type 1b

Blood ◽  
2003 ◽  
Vol 101 (6) ◽  
pp. 2381-2387 ◽  
Author(s):  
Rosanna Leuzzi ◽  
Gábor Bánhegyi ◽  
Tamás Kardon ◽  
Paola Marcolongo ◽  
Piero-Leopoldo Capecchi ◽  
...  
Keyword(s):  
Glycogen Storage Disease ◽  
Storage Disease ◽  
Glycogen Storage Disease Type ◽  
Phosphate Transport ◽  
Glycogen Storage ◽  
Disease Type ◽  
Jurkat Cells ◽  
Phosphate Transporter ◽  
Disulfonic Acid ◽  
Human Neutrophils

Mutations in the gene of the hepatic glucose-6-phosphate transporter cause glycogen storage disease type 1b. In this disease, the altered glucose homeostasis and liver functions are accompanied by an impairment of neutrophils/monocytes. However, neither the existence of a microsomal glucose-6-phosphate transport, nor the connection between its defect and cell dysfunction has been demonstrated in neutrophils/monocytes. In this study we have characterized the microsomal glucose-6-phosphate transport of human neutrophils and differentiated HL-60 cells. The transport of glucose-6-phosphate was sensitive to the chlorogenic acid derivative S3483,N-ethylmaleimide, and 4,4′-diisothiocyanostilbene-2,2′-disulfonic acid, known inhibitors of the hepatic microsomal glucose-6-phosphate transporter. A glucose-6-phosphate uptake was also present in microsomes from undifferentiated HL-60 and Jurkat cells, but it was insensitive to S3483. The treatment with S3484 of intact human neutrophils and differentiated HL-60 cells mimicked some leukocyte defects of glycogen storage disease type 1b patients (ie, the drug inhibited phorbol myristate acetate–induced superoxide anion production and reduced the size of endoplasmic reticulum Ca2+ stores). Importantly, the treatment with S3484 also resulted in apoptosis of human neutrophils and differentiated HL-60 cells, while undifferentiated HL-60 and Jurkat cells were unaffected by the drug. The proapoptotic effect of S3483 was prevented by the inhibition of nicotinamide adenine dinucleotide phosphate oxidase or by antioxidant treatment. These results suggest that microsomal glucose-6-phosphate transport has a role in the antioxidant protection of neutrophils, and that the genetic defect of the transporter leads to the impairment of cellular functions and apoptosis.

scholarly journals A Phosphate Transporter from the Root Endophytic FungusPiriformospora indicaPlays a Role in Phosphate Transport to the Host Plant

2010 ◽  
Vol 285 (34) ◽  
pp. 26532-26544 ◽  
Author(s):  
Vikas Yadav ◽  
Manoj Kumar ◽  
Deepak Kumar Deep ◽  
Hemant Kumar ◽  
Ruby Sharma ◽  
...  
Keyword(s):  
Host Plant ◽  
Phosphate Transport ◽  
Phosphate Transporter

scholarly journals Role of coiled-coil registry shifts in activation of human Bicaudal D2 for dynein recruitment upon cargo-binding

2019 ◽  
Author(s):  
Crystal R. Noell ◽  
Jia Ying Loh ◽  
Erik W. Debler ◽  
Kyle M. Loftus ◽  
Heying Cui ◽  
...  
Keyword(s):  
Muscle Development ◽  
Muscular Atrophy ◽  
Coiled Coil ◽  
Coiled Coils ◽  
General Mechanism ◽  
List Type ◽  
Transport Pathways ◽  
Dynamics Simulations ◽  
Dependent Transport ◽  
Cargo Binding

SUMMARYDynein adaptors such as Bicaudal D2 (BicD2) recognize cargo for dynein-dependent transport. BicD2-dependent transport pathways are important for brain and muscle development. Cargo-bound adaptors are required to activate dynein for processive transport, but the mechanism of action is elusive. Here, we report the structure of the cargo-binding domain of human BicD2 that forms a dimeric coiled-coil with homotypic registry, in which both helices are aligned. To investigate if BicD2 can switch to an asymmetric registry, where a portion of one helix is vertically shifted, we performed molecular dynamics simulations. Both registry types are stabilized by distinct conformations of F743. For the F743I variant, which increases dynein recruitment in the Drosophila homolog, and for the human R747C variant, which causes spinal muscular atrophy, spontaneous coiled-coil registry shifts are observed, which may cause the BicD2-hyperactivation phenotype and disease. We propose that a registry shift upon cargo-binding activates auto-inhibited BicD2 for dynein recruitment.HighlightsStable, bona fide BicD2 coiled-coils with distinct registries can be formed.We provide evidence that a human disease mutation causes a coiled-coil registry shift.A coiled-coil registry shift could relieve BicD2-autoinhibition upon cargo-binding.The ability to undergo registry shifts may be an inherent property of coiled-coils.In BriefOur results support that stable coiled-coils of BicD2 with distinct registries can be formed, and suggest a molecular mechanism for such registry switches. We provide evidence that disease-causing mutations in coiled-coils may alter the equilibrium between registry-shifted conformers, which we propose as a general mechanism of pathogenesis for coiled-coils.Graphical Abstract

scholarly journals The shape of the bacterial ribosome exit tunnel affects cotranslational protein folding

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Renuka Kudva ◽  
Pengfei Tian ◽  
Fátima Pardo-Avila ◽  
Marta Carroni ◽  
Robert B Best ◽  
...  
Keyword(s):  
Protein Folding ◽  
Protein Domain ◽  
Coarse Grained ◽  
E Coli ◽  
Folded Structure ◽  
Bacterial Ribosome ◽  
Folded Proteins ◽  
Exit Tunnel ◽  
Ribosome Exit Tunnel ◽  
Dynamics Simulations

The E. coli ribosome exit tunnel can accommodate small folded proteins, while larger ones fold outside. It remains unclear, however, to what extent the geometry of the tunnel influences protein folding. Here, using E. coli ribosomes with deletions in loops in proteins uL23 and uL24 that protrude into the tunnel, we investigate how tunnel geometry determines where proteins of different sizes fold. We find that a 29-residue zinc-finger domain normally folding close to the uL23 loop folds deeper in the tunnel in uL23 Δloop ribosomes, while two ~ 100 residue proteins normally folding close to the uL24 loop near the tunnel exit port fold at deeper locations in uL24 Δloop ribosomes, in good agreement with results obtained by coarse-grained molecular dynamics simulations. This supports the idea that cotranslational folding commences once a protein domain reaches a location in the exit tunnel where there is sufficient space to house the folded structure.

scholarly journals Mfsd2a and Spns2 are essential for sphingosine-1-phosphate transport in the formation and maintenance of the blood-brain barrier

2020 ◽  
Vol 6 (22) ◽  
pp. eaay8627 ◽  
Author(s):  
Zhifu Wang ◽  
Yongtao Zheng ◽  
Fan Wang ◽  
Junjie Zhong ◽  
Tong Zhao ◽  
...  
Keyword(s):  
Blood Brain Barrier ◽  
Phosphate Transport ◽  
Major Facilitator Superfamily ◽  
Brain Barrier ◽  
Specific Marker ◽  
Blood Brain ◽  
Sphingosine 1 Phosphate ◽  
The Central Nervous System ◽  
Major Facilitator ◽  
Brain Homeostasis

To maintain brain homeostasis, a unique interface known as the blood-brain barrier (BBB) is formed between the blood circulation and the central nervous system (CNS). Major facilitator superfamily domain-containing 2a (Mfsd2a) is a specific marker of the BBB. However, the mechanism by which Mfsd2a influences the BBB is poorly understood. In this study, we demonstrated that Mfsd2a is essential for sphingosine-1-phosphate (S1P) export from endothelial cells in the brain. We found that Mfsd2a and Spinster homolog 2 (Spns2) form a protein complex to ensure the efficient transport of S1P. Furthermore, the S1P-rich microenvironment in the extracellular matrix (ECM) in the vascular endothelium dominates the formation and maintenance of the BBB. We demonstrated that different concentrations of S1P have different effects on BBB integrity. These findings help to unravel the mechanism by which S1P regulates BBB and also provide previously unidentified insights into the delivery of neurological drugs in the CNS.

Phosphate Transport in Epithelial and Nonepithelial Tissue

2021 ◽  
Vol 101 (1) ◽  
pp. 1-35 ◽  
Author(s):  
Nati Hernando ◽  
Kenneth Gagnon ◽  
Eleanor Lederer
Keyword(s):  
Intracellular Transport ◽  
Bone Mineralization ◽  
Helical Structure ◽  
Serum Levels ◽  
Tissue Expression ◽  
Phosphate Transport ◽  
High Energy ◽  
Normal Serum ◽  
Phosphate Transporters ◽  
The Individual

Phosphate is an essential nutrient for life and is a critical component of bone formation, a major signaling molecule, and structural component of cell walls. Phosphate is also a component of high-energy compounds (i.e., AMP, ADP, and ATP) and essential for nucleic acid helical structure (i.e., RNA and DNA). Phosphate plays a central role in the process of mineralization, normal serum levels being associated with appropriate bone mineralization, while high and low serum levels are associated with soft tissue calcification. The serum concentration of phosphate and the total body content of phosphate are highly regulated, a process that is accomplished by the coordinated effort of two families of sodium-dependent transporter proteins. The three isoforms of the SLC34 family (SLC34A1–A3) show very restricted tissue expression and regulate intestinal absorption and renal excretion of phosphate. SLC34A2 also regulates the phosphate concentration in multiple lumen fluids including milk, saliva, pancreatic fluid, and surfactant. Both isoforms of the SLC20 family exhibit ubiquitous expression (with some variation as to which one or both are expressed), are regulated by ambient phosphate, and likely serve the phosphate needs of the individual cell. These proteins exhibit similarities to phosphate transporters in nonmammalian organisms. The proteins are nonredundant as mutations in each yield unique clinical presentations. Further research is essential to understand the function, regulation, and coordination of the various phosphate transporters, both the ones described in this review and the phosphate transporters involved in intracellular transport.

Sign in / Sign up

Export Citation Format

Share Document