Solute Transport through Mitochondrial Porins In Vitro and In Vivo.

Mitochondria are most likely descendants of strictly aerobic prokaryotes from the class . The mitochondrial matrix is surrounded by two membranes according to its relationship with Gram-negative bacteria. Similar to the bacterial outer membrane, the mitochondrial outer membrane acts as a molecular sieve because it also contains diffusion pores.

Unconventional structure and mechanisms for membrane interaction and translocation of the NF-κB-targeting toxin AIP56.

Bacterial AB toxins are secreted key virulence factors that are internalized by target cells through receptor-mediated endocytosis, translocating their enzymatic domain to the cytosol from endosomes (short-trip) or the endoplasmic reticulum (long-trip). To accomplish this, bacterial AB toxins evolved a multidomain structure organized into either a single polypeptide chain or non-covalently associated polypeptide chains. The prototypical short-trip single-chain toxin is characterized by a receptor-binding domain that confers cellular specificity and a translocation domain responsible for pore formation whereby the catalytic domain translocates to the cytosol in an endosomal acidification-dependent way.

Chloroquine-analogues block anthrax protective antigen channels in steady-state and kinetic studies.

The tripartite anthrax toxin from Bacillus anthracis represents the prototype of A-B type of toxins, where the effector A (an enzymatic subunit) is transported with the help of a binding component B into a target cell. Anthrax toxin consists of three different molecules, two effectors, lethal factor (LF) and edema factor (EF) and the binding component also known as protective antigen (PA). PA forms heptamers or octamers following binding to host cell's receptors and mediates the translocation of the effectors into the cytosol via the endosomal pathway.

Conformational Dynamics of Loop L3 in OmpF: Implications toward Antibiotic Translocation and Voltage Gating.

In the present work, we delineate the molecular mechanism of a bulky antibiotic permeating through a bacterial channel and uncover the role of conformational dynamics of the constriction loop in this process. Using the temperature accelerated sliced sampling approach, we shed light onto the dynamics of the L3 loop, in particular the F118 to S125 segment, at the constriction regions of the OmpF porin. We complement the findings with single channel electrophysiology experiments and applied-field simulations, and we demonstrate the role of hydrogen-bond stabilization in the conformational dynamics of the L3 loop.

Importance of the lysine cluster in the translocation of anions through the pyrophosphate specific channel OprO.

Pseudomonas aeruginosa is a Gram-negative bacterium with an intrinsic resistance towards antibiotics due to the lack of a large diffusion pores. Exchange of substances with the environment is done mainly through a set of narrow and substrate-specific porins in its outer membrane that filter molecules according to their size and chemical composition. Among these proteins are OprP and OprO involved in the selective uptake of mono- and pyrophosphates, respectively.

Cell wall channels of Rhodococcus species: identification and characterization of the cell wall channels of Rhodococcus corynebacteroides and Rhodococcus ruber.

The cell wall of Rhodococcus corynebacteroides formerly known as Nocardia corynebacteroides contains cell wall channels that are responsible for the cell wall permeability of this bacterium. Based on partial sequencing of the polypeptide subunits and a BLAST search, we identified one polypeptide of R. corynebacteroides (PorARc) and two polypeptides (PorARr and PorBRr) from the closely related bacterium Rhodococcus ruber.

Historical Perspective of Pore-Forming Activity Studies of Voltage-Dependent Anion Channel (Eukaryotic or Mitochondrial Porin) Since Its Discovery in the 70th of the Last Century.

Eukaryotic porin, also known as Voltage-Dependent Anion Channel (VDAC), is the most frequent protein in the outer membrane of mitochondria that are responsible for cellular respiration. Mitochondria are most likely descendants of strictly aerobic Gram-negative bacteria from the α-proteobacterial lineage. In accordance with the presumed ancestor, mitochondria are surrounded by two membranes.

Structure and Gating Behavior of the Human Integral Membrane Protein VDAC1 in a Lipid Bilayer.

The voltage-dependent anion channel (VDAC), the most abundant protein in the outer mitochondrial membrane, is responsible for the transport of all ions and metabolites into and out of mitochondria. Larger than any of the β-barrel structures determined to date by magic-angle spinning (MAS) NMR, but smaller than the size limit of cryo-electron microscopy (cryo-EM), VDAC1's 31 kDa size has long been a bottleneck in determining its structure in a near-native lipid bilayer environment. Using a single two-dimensional (2D) crystalline sample of human VDAC1 in lipids, we applied proton-detected fast magic-angle spinning NMR spectroscopy to determine the arrangement of β strands.

Permeation of Fosfomycin through the Phosphate-Specific Channels OprP and OprO of .

is a Gram-negative opportunistic pathogen responsible for many nosocomial infections. It is quite resistant to various antibiotics, caused by the absence of general diffusion pores in the outer membrane. Instead, it contains many substrate-specific channels.

Clostridium perfringens Beta2 toxin forms highly cation-selective channels in lipid bilayers.

Clostridium perfringens is a potent producer of a variety of toxins. Well studied from these are five toxins (alpha, Beta (CPB), epsilon, iota and CPE) that are produced by seven toxinotype strains (A-G) of C. perfringens.

Characterization and Pharmacological Inhibition of the Pore-Forming CDTb Toxin.

The clinically highly relevant releases several AB-type toxins that cause diseases such as diarrhea and pseudomembranous colitis. In addition to the main virulence factors Rho/Ras-glycosylating toxins TcdA and TcdB, hypervirulent strains produce the binary AB-type toxin CDT. CDT consists of two separate proteins.

Fosmidomycin transport through the phosphate-specific porins OprO and OprP of Pseudomonas aeruginosa.

The Gram-negative bacterium Pseudomonas aeruginosa is an opportunistic pathogen, responsible for many hospital-acquired infections. The bacterium is quite resistant toward many antibiotics, in particular because of the fine-tuned permeability of its outer membrane (OM). General diffusion outer membrane pores are quite rare in this organism.

Characteristics of the Protein Complexes and Pores Formed by Hemolysin BL.

Hemolysin BL is a tripartite toxin responsible for a diarrheal type of food poisoning. Open questions remain regarding its mode of action, including the extent to which complex formation prior to cell binding contributes to pore-forming activity, how these complexes are composed, and the properties of the pores formed in the target cell membrane. Distinct complexes of up to 600 kDa were found on native gels, whose structure and size were primarily defined by Hbl B.

RTX-Toxins.

RTX-Toxins (Repeats in ToXin) are members of a rapidly expanding family of proteins [...

Central residues of the amphipathic β-hairpin loop control the properties of Clostridium perfringens epsilon-toxin channel.

Clostridium perfringens epsilon toxin (ETX) is a heptameric pore-forming toxin of the aerolysin toxin family. ETX is the most potent toxin of this toxin family and the third most potent bacterial toxin with high cytotoxic and lethal activities in animals. In addition, ETX shows a demyelinating activity in nervous tissue leading to devastating multifocal central nervous system white matter disease in ruminant animals.

Human peptide α-defensin-1 interferes with Clostridioides difficile toxins TcdA, TcdB, and CDT.

The human pathogenic bacterium Clostridioides difficile produces two exotoxins TcdA and TcdB, which inactivate Rho GTPases thereby causing C. difficile-associated diseases (CDAD) including life-threatening pseudomembranous colitis. Hypervirulent strains produce additionally the binary actin ADP-ribosylating toxin CDT.

Membrane Activity and Channel Formation of the Adenylate Cyclase Toxin (CyaA) of in Lipid Bilayer Membranes.

The Gram-negative bacterium is the cause of whooping cough. One of its pathogenicity factors is the adenylate cyclase toxin (CyaA) secreted by a Type I export system. The 1706 amino acid long CyaA (177 kDa) belongs to the continuously increasing family of repeat in toxin (RTX) toxins because it contains in its C-terminal half a high number of nine-residue tandem repeats.

LtxA Hijacks Endocytic Trafficking Pathways in Human Lymphocytes.

Leukotoxin (LtxA), from oral pathogen is a secreted membrane-damaging protein. LtxA is internalized by β2 integrin LFA-1 (CD11a/CD18)-expressing leukocytes and ultimately causes cell death; however, toxin localization in the host cell is poorly understood and these studies fill this void. We investigated LtxA trafficking using multi-fluor confocal imaging, flow cytometry and Rab5a knockdown in human T lymphocyte Jurkat cells.

Structure of a Tc holotoxin pore provides insights into the translocation mechanism.

Tc toxins are modular toxin systems of insect and human pathogenic bacteria. They are composed of a 1.4-MDa pentameric membrane translocator (TcA) and a 250-kDa cocoon (TcB and TcC) encapsulating the 30-kDa toxic enzyme (C terminus of TcC).

Channel Formation by LktA of in Lipid Bilayer Membranes and Comparison of Channel Properties with Other RTX-Cytolysins.

Cytolysin LktA is one of the major pathogenicity factors of (formerly ) that is the cause of pasteurellosis, also known as shipping fever pneumonia, causing substantial loss of sheep and cattle during transport. LktA belongs to the family of RTX-toxins (Repeats in ToXins) that are produced as pathogenicity factors by a variety of Gram-negative bacteria. Sublytic concentrations of LktA cause inflammatory responses of ovine leukocytes.

Role of AIP56 disulphide bond and its reduction by cytosolic redox systems for efficient intoxication.

Apoptosis-inducing protein of 56 kDa (AIP56) is a major virulence factor of Photobacterium damselae subsp. piscicida, a gram-negative pathogen that infects warm water fish species worldwide and causes serious economic losses in aquacultures. AIP56 is a single-chain AB toxin composed by two domains connected by an unstructured linker peptide flanked by two cysteine residues that form a disulphide bond.

Revisiting an old antibiotic: bacitracin neutralizes binary bacterial toxins and protects cells from intoxication.

The antibiotic bacitracin (Bac) inhibits cell wall synthesis of gram-positive bacteria. Here, we discovered a totally different activity of Bac: the neutralization of bacterial exotoxins. Bac prevented intoxication of mammalian cells with the binary enterotoxins Clostridium botulinum C2, C.

Enterotoxin: The Toxin Forms Highly Cation-Selective Channels in Lipid Bilayers.

One of the numerous toxins produced by is enterotoxin (CPE), a polypeptide with a molecular mass of 35.5 kDa exhibiting three different domains. Domain one is responsible for receptor binding, domain two is involved in hexamer formation and domain three has to do with channel formation in membranes.

Generation of a recombinant Aggregatibacter actinomycetemcomitans RTX toxin in Escherichia coli.

A leukotoxin (LtxA) that is produced by Aggregatibacter actinomycetemcomitans (Aa) is an important virulence determinant in an aggressive form of periodontitis in adolescents. Understanding the function of this protein at the molecular level is critical to elucidating its role in the disease process. To accomplish genetic analysis of the protein structure and relating these observations to toxin function, we have developed an E.

The major outer membrane protein of Legionella pneumophila Lpg1974 shows pore-forming characteristics similar to the human mitochondrial outer membrane pore, hVDAC1.

Legionella pneumophila is an aerobic and nonspore-forming pathogenic Gram-negative bacterium of the genus Legionella. It is the causative agent of Legionnaires' disease, also known as Legionellosis. The hosts of this organism are diverse, ranging from simple water borne protozoans such as amoebae to more complex hosts such as macrophages in humans.