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Biomolecules Jan 2021Cell plasma membrane proteins are considered as gatekeepers of the cell and play a major role in regulating various processes. Transport proteins constitute a subclass... (Review)
Review
Cell plasma membrane proteins are considered as gatekeepers of the cell and play a major role in regulating various processes. Transport proteins constitute a subclass of cell plasma membrane proteins enabling the exchange of molecules and ions between the extracellular environment and the cytosol. A plethora of human pathologies are associated with the altered expression or dysfunction of cell plasma membrane transport proteins, making them interesting therapeutic drug targets. However, the search for therapeutics is challenging, since many drug candidates targeting cell plasma membrane proteins fail in (pre)clinical testing due to inadequate selectivity, specificity, potency or stability. These latter characteristics are met by nanobodies, which potentially renders them eligible therapeutics targeting cell plasma membrane proteins. Therefore, a therapeutic nanobody-based strategy seems a valid approach to target and modulate the activity of cell plasma membrane transport proteins. This review paper focuses on methodologies to generate cell plasma membrane transport protein-targeting nanobodies, and the advantages and pitfalls while generating these small antibody-derivatives, and discusses several therapeutic nanobodies directed towards transmembrane proteins, including channels and pores, adenosine triphosphate-powered pumps and porters.
Topics: Antigens; Cell Membrane; Humans; Membrane Transport Proteins; Models, Biological; Single-Domain Antibodies
PubMed: 33418902
DOI: 10.3390/biom11010063 -
The Protein Journal Aug 2019The twin-arginine protein translocation (Tat) system has been characterized in bacteria, archaea and the chloroplast thylakoidal membrane. This system is distinct from... (Review)
Review
The twin-arginine protein translocation (Tat) system has been characterized in bacteria, archaea and the chloroplast thylakoidal membrane. This system is distinct from other protein transport systems with respect to two key features. Firstly, it accepts cargo proteins with an N-terminal signal peptide that carries the canonical twin-arginine motif, which is essential for transport. Second, the Tat system only accepts and translocates fully folded cargo proteins across the respective membrane. Here, we review the core essential features of folded protein transport via the bacterial Tat system, using the three-component TatABC system of Escherichia coli and the two-component TatAC systems of Bacillus subtilis as the main examples. In particular, we address features of twin-arginine signal peptides, the essential Tat components and how they assemble into different complexes, mechanistic features and energetics of Tat-dependent protein translocation, cytoplasmic chaperoning of Tat cargo proteins, and the remarkable proofreading capabilities of the Tat system. In doing so, we present the current state of our understanding of Tat-dependent protein translocation across biological membranes, which may serve as a lead for future investigations.
Topics: Arginine; Bacillus subtilis; Cell Membrane; Escherichia coli; Escherichia coli Proteins; Membrane Transport Proteins; Protein Folding; Protein Sorting Signals; Protein Transport; SEC Translocation Channels; Twin-Arginine-Translocation System
PubMed: 31401776
DOI: 10.1007/s10930-019-09859-y -
Current Opinion in Structural Biology Aug 2019The Golgi apparatus plays a central role in the secretory pathway as a hub for posttranslational modification, protein sorting and quality control. To date, there is... (Review)
Review
The Golgi apparatus plays a central role in the secretory pathway as a hub for posttranslational modification, protein sorting and quality control. To date, there is little structural or biochemical information concerning the function of transporters that reside within this organelle. The SLC35 family of nucleotide sugar transporters link the synthesis of activated sugar molecules and sulfate in the cytoplasm, with the luminal transferases that catalyse their attachment to proteins and lipids during glycosylation and sulfation. A recent crystal structure of the GDP-mannose transporter has revealed key sequence motifs that direct ligand recognition and transport. Further biochemical studies unexpectedly found a requirement for short chain lipids in activating the transporter, suggesting a possible route for transport regulation within the Golgi.
Topics: Animals; Golgi Apparatus; Humans; Membrane Transport Proteins; Nucleotides; Sugars
PubMed: 30999236
DOI: 10.1016/j.sbi.2019.03.019 -
International Journal of Molecular... Jul 2022Ultrashort peptides (USPs), consisting of 2-7 amino-acid residues, are a group of signaling molecules that regulate gene expression and protein synthesis under normal... (Review)
Review
Ultrashort peptides (USPs), consisting of 2-7 amino-acid residues, are a group of signaling molecules that regulate gene expression and protein synthesis under normal conditions in various diseases and ageing. USPs serve as a basis for the development of drugs with a targeted mechanism of action. The purpose of this review is to systematize the available data on USP transport involving POT and LAT transporters in various organs and tissues under normal, pathological and ageing conditions. The carriers of the POT family (PEPT1, PEPT2, PHT1, PHT2) transport predominantly di- and tripeptides into the cell. Methods of molecular modeling and physicochemistry have demonstrated the ability of LAT1 to transfer not only amino acids but also some di- and tripeptides into the cell and out of it. LAT1 and 2 are involved in the regulation of the antioxidant, endocrine, immune and nervous systems' functions. Analysis of the above data allows us to conclude that, depending on their structure, di- and tripeptides can be transported into the cells of various tissues by POT and LAT transporters. This mechanism is likely to underlie the tissue specificity of peptides, their geroprotective action and effectiveness in the case of neuroimmunoendocrine system disorders.
Topics: Amino Acids; Biological Transport; Membrane Transport Proteins; Organ Specificity; Peptides; Symporters
PubMed: 35887081
DOI: 10.3390/ijms23147733 -
Journal of Cell Science Jul 2023Mitochondrial protein import is essential for organellar biogenesis, and thereby for the sufficient supply of cytosolic ATP - which is particularly important for cells...
Mitochondrial protein import is essential for organellar biogenesis, and thereby for the sufficient supply of cytosolic ATP - which is particularly important for cells with high energy demands like neurons. This study explores the prospect of import machinery perturbation as a cause of neurodegeneration instigated by the accumulation of aggregating proteins linked to disease. We found that the aggregation-prone Tau variant (TauP301L) reduces the levels of components of the import machinery of the outer (TOM20, encoded by TOMM20) and inner membrane (TIM23, encoded by TIMM23) while associating with TOM40 (TOMM40). Intriguingly, this interaction affects mitochondrial morphology, but not protein import or respiratory function; raising the prospect of an intrinsic rescue mechanism. Indeed, TauP301L induced the formation of tunnelling nanotubes (TNTs), potentially for the recruitment of healthy mitochondria from neighbouring cells and/or the disposal of mitochondria incapacitated by aggregated Tau. Consistent with this, inhibition of TNT formation (and rescue) reveals Tau-induced import impairment. In primary neuronal cultures, TauP301L induced morphological changes characteristic of neurodegeneration. Interestingly, these effects were mirrored in cells where the import sites were blocked artificially. Our results reveal a link between aggregation-prone Tau and defective mitochondrial import relevant to disease.
Topics: Mitochondria; Membrane Transport Proteins; Protein Transport; Receptors, Cell Surface; Neurons; Mitochondrial Proteins; Mitochondrial Membrane Transport Proteins
PubMed: 37303235
DOI: 10.1242/jcs.260993 -
The EMBO Journal Aug 2023Efflux of antibacterial compounds is a major mechanism for developing antimicrobial resistance. In the Gram-positive pathogen Staphylococcus aureus, QacA, a 14...
Efflux of antibacterial compounds is a major mechanism for developing antimicrobial resistance. In the Gram-positive pathogen Staphylococcus aureus, QacA, a 14 transmembrane helix containing major facilitator superfamily antiporter, mediates proton-coupled efflux of mono and divalent cationic antibacterial compounds. In this study, we report the cryo-EM structure of QacA, with a single mutation D411N that improves homogeneity and retains efflux activity against divalent cationic compounds like dequalinium and chlorhexidine. The structure of substrate-free QacA, complexed to two single-domain camelid antibodies, was elucidated to a resolution of 3.6 Å. The structure displays an outward-open conformation with an extracellular helical hairpin loop (EL7) between transmembrane helices 13 and 14, which is conserved in a subset of DHA2 transporters. Removal of the EL7 hairpin loop or disrupting the interface formed between EL7 and EL1 compromises efflux activity. Chimeric constructs of QacA with a helical hairpin and EL1 grafted from other DHA2 members, LfrA and SmvA, restore activity in the EL7 deleted QacA revealing the allosteric and vital role of EL7 hairpin in antibacterial efflux in QacA and related members.
Topics: Humans; Staphylococcus aureus; Cryoelectron Microscopy; Bacterial Proteins; Membrane Transport Proteins; Anti-Bacterial Agents; Staphylococcal Infections
PubMed: 37458117
DOI: 10.15252/embj.2023113418 -
Biochemistry Nov 2021Iron is an essential nutrient for virtually every living organism, especially pathogenic prokaryotes. Despite its importance, however, both the acquisition and the... (Review)
Review
Iron is an essential nutrient for virtually every living organism, especially pathogenic prokaryotes. Despite its importance, however, both the acquisition and the export of this element require dedicated pathways that are dependent on oxidation state. Due to its solubility and kinetic lability, reduced ferrous iron (Fe) is useful to bacteria for import, chaperoning, and efflux. Once imported, ferrous iron may be loaded into apo and nascent enzymes and even sequestered into storage proteins under certain conditions. However, excess labile ferrous iron can impart toxicity as it may spuriously catalyze Fenton chemistry, thereby generating reactive oxygen species and leading to cellular damage. In response, it is becoming increasingly evident that bacteria have evolved Fe efflux pumps to deal with conditions of ferrous iron excess and to prevent intracellular oxidative stress. In this work, we highlight recent structural and mechanistic advancements in our understanding of prokaryotic ferrous iron import and export systems, with a focus on the connection of these essential transport systems to pathogenesis. Given the connection of these pathways to the virulence of many increasingly antibiotic resistant bacterial strains, a greater understanding of the mechanistic details of ferrous iron cycling in pathogens could illuminate new pathways for future therapeutic developments.
Topics: Anti-Bacterial Agents; Bacteria; Biological Transport; Catalysis; Homeostasis; Ion Transport; Iron; Kinetics; Membrane Proteins; Membrane Transport Proteins; Oxidation-Reduction; Oxidative Stress; Prokaryotic Cells; Reactive Oxygen Species; Solubility; Virulence
PubMed: 34670078
DOI: 10.1021/acs.biochem.1c00586 -
Antimicrobial Agents and Chemotherapy Oct 2023Drug-resistant is a worldwide health-care problem rendering current tuberculosis (TB) drugs ineffective. Drug efflux is an important mechanism in bacterial drug...
Drug-resistant is a worldwide health-care problem rendering current tuberculosis (TB) drugs ineffective. Drug efflux is an important mechanism in bacterial drug resistance. The MmpL4 and MmpL5 transporters form functionally redundant complexes with their associated MmpS4 and MmpS5 proteins and constitute the inner membrane components of an essential siderophore secretion system of . Inactivating siderophore secretion is toxic for due to self-poisoning at low-iron conditions and leads to a strong virulence defect in mice. In this study, we show that mutants lacking components of the MmpS4-MmpL4 and MmpS5-MmpL5 systems are more susceptible to bedaquiline, clofazimine, and rifabutin, important drugs for treatment of drug-resistant TB. While genetic deletion experiments revealed similar functions of the MmpL4 and MmpL5 transporters in siderophore and drug secretion, complementation experiments indicated that the MmpS4-MmpL4 proteins alone are not sufficient to restore drug efflux in an mutant lacking both operons, in contrast to MmpS5-MmpL5. Importantly, an mutant lacking the recently discovered periplasmic Rv0455c protein, which is also essential for siderophore secretion, is more susceptible to the same drugs. These results reveal a promising target for the development of dual-function TB drugs, which might poison by blocking siderophore secretion and synergize with other drugs by impairing drug efflux.
Topics: Animals; Mice; Mycobacterium tuberculosis; Siderophores; Tuberculosis; Membrane Transport Proteins; Tuberculosis, Multidrug-Resistant; Antitubercular Agents
PubMed: 37676015
DOI: 10.1128/aac.01629-22 -
Current Opinion in Microbiology Jun 2021All mechanisms of clinical antibiotic resistance benefit from activities of polyspecific efflux pumps acting to reduce intracellular accumulation of toxins and... (Review)
Review
All mechanisms of clinical antibiotic resistance benefit from activities of polyspecific efflux pumps acting to reduce intracellular accumulation of toxins and antibiotics. In Gram-negative bacteria, the major polyspecific efflux transporters belong to the Resistance-Nodulation-cell Division (RND) superfamily of proteins, which are capable of expelling thousands of structurally diverse compounds. Recent structural and functional advances generated novel insights into mechanisms underlying the biochemical versatility of RND transporters. This opinion article reviews these mechanisms and discusses implications of the polyspecificity of RND transporters for bacterial survival and for the development of efflux pump inhibitors effective in clinics.
Topics: Anti-Bacterial Agents; Bacterial Proteins; Biological Transport; Drug Resistance, Microbial; Drug Resistance, Multiple, Bacterial; Gram-Negative Bacteria; Membrane Transport Proteins
PubMed: 33940284
DOI: 10.1016/j.mib.2021.03.009 -
NMR in Biomedicine Jan 2021X-linked creatine transporter deficiency (CTD) is one of the three types of cerebral creatine deficiency disorders. CTD arises from pathogenic variants in the X-linked...
X-linked creatine transporter deficiency (CTD) is one of the three types of cerebral creatine deficiency disorders. CTD arises from pathogenic variants in the X-linked gene SLC6A8. We report the first phosphorus ( P) MRS study of patients with CTD, where both phosphocreatine and total creatine concentrations were found to be markedly reduced. Despite the diminished role of creatine and phosphocreatine in oxidative phosphorylation in CTD, we found no elevation of lactate or lowered pH, indicating that the brain energy supply still largely relied on oxidative metabolism. Our results suggest that mitochondrial function is a potential therapeutic target for CTD.
Topics: Adolescent; Brain; Child; Child, Preschool; Creatine; Humans; Magnetic Resonance Imaging; Male; Membrane Transport Proteins; Metabolome; Oxidative Phosphorylation; Phosphorus; Proton Magnetic Resonance Spectroscopy
PubMed: 32990357
DOI: 10.1002/nbm.4419