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Kaitlynne Bohm, Steffen Porwollik, Weiping Chu, John A Dover, Eddie B Gilcrease, Sherwood R Casjens, Michael McClelland, Kristin N Parent
Bacteriophages rely on their hosts for replication, and many host genes critically determine either viral progeny production or host success via phage resistance. A random insertion transposon library of 240,000 mutants in Salmonella enterica serovar Typhimurium was used to monitor effects of individual bacterial gene disruptions on bacteriophage P22 lytic infection. These experiments revealed candidate host genes that alter the timing of phage P22 propagation. Using a False Discovery Rate of <0.1, mutations in 235 host genes either blocked or delayed progression of P22 lytic infection, including many genes for which this role was previously unknown...
February 22, 2018: Molecular Microbiology
Yu-Cheng Lin, Matthew D Sekedat, William Cole Cornell, Gustavo M Silva, Chinweike Okegbe, Alexa Price-Whelan, Christine Vogel, Lars E P Dietrich
Microbes in biofilms face the challenge of substrate limitation. In particular, cells in Pseudomonas aeruginosa biofilms growing in the laboratory or during host colonization often become limited for oxygen. Previously we found that phenazines, antibiotics produced by P. aeruginosa , balance the intracellular redox state for cells in biofilms. Here, we show that genes involved in denitrification are induced in phenazine-null (Δ phz ) mutant biofilms grown under an aerobic atmosphere, even in the absence of nitrate...
February 20, 2018: Journal of Bacteriology
María J Torres, Sergio Avila, Eulogio J Bedmar, María J Delgado
The alfalfa endosymbiont Ensifer meliloti strain1021 is known to be an incomplete denitrifier due to its inability to grow anoxically using nitrate as respiratory substrate to produce ATP and grow under anoxic conditions. Although this bacterium contains and expresses the complete set of denitrification genes napEFDABC, nirK, norECBQD and nosRZDFYLX encoding the periplasmic nitrate reductase (Nap), Cu-containing nitrite reductase (NirK), c-type nitric oxide (cNor) and nitrous oxide reductase (Nos), respectively, the reasons of its inability to grow under anoxic conditions are still very poorly understood...
February 16, 2018: FEMS Microbiology Letters
Mateusz Kędzior, Aleksandra Pawlak, Rafał Seredyński, Jacek Bania, Aleksandra Platt-Samoraj, Magdalena Czemplik, Elżbieta Klausa, Gabriela Bugla-Płoskońska, Jan Gutowicz
Cysteine proteases of the papain family, including mammalian cathepsins, play important physiological roles, however, their excessive activity may contribute to the development of various pathologies. Therefore, cysteine cathepsin inhibitors are being considered as promising drugs to treat cathepsin-driven diseases. Diverse saprophytic and parasitic microbes produce such inhibitors, which target the host's proteases playing pivotal roles in immune responses, thus leading to the survival of microbes within their host...
March 2018: Microbiological Research
Abdelrahim Zoued, Jean-Pierre Duneau, Eric Durand, Alexandre P España, Laure Journet, Françoise Guerlesquin, Eric Cascales
The Type VI secretion system (T6SS) is a multiprotein complex used by bacteria to deliver effectors into target cells. The T6SS comprises a bacteriophage-like contractile tail structure anchored to the cell envelope by a membrane complex constituted of the TssJ outer membrane lipoprotein and the TssL and TssM inner membrane proteins. TssJ establishes contact with the periplasmic domain of TssM whereas the transmembrane segments of TssM and its cytoplasmic domain interact with TssL. TssL protrudes in the cytoplasm but is anchored by a C-terminal transmembrane helix (TMH)...
February 16, 2018: Journal of Molecular Biology
Mohammad Reza Bagherinejad, Hamid Mir-Mohammad Sadeghi, Daryoush Abedi, Fateme Moazen, Mohammad Rabbani
Background: Twine-arginine translocation (TAT) system is one of the exporting systems in Escherichia coli which could transport fully/semi-correctly folded proteins outside the reductive cytoplasmic space. In combination with co-expression with a chaperone system, the correctly folded proteins could be transported to oxidative periplasmic space and culture media to pass the main limitations in E. coli expression system such as misfolding and inclusion body formation. Materials and Methods: To study the effectiveness of signaling sequences and chaperone co-expression on the translocation of expressed protein, somatropin was selected as the target...
2018: Advanced Biomedical Research
Felipe Cava
The L-forms of amino acids are used in all kingdoms of life to synthesize proteins. However, the bacterium Vibriocholerae, the causative agent of cholera, produces D-amino acids which are released to the environment at millimolar concentrations. We baptized these D-amino acids as non-canonical D-amino acids (NCDAAs) since they are different from those (i.e. D-alanine and D-glutamate) normally present in the bacterial cell wall. In V. cholerae, production of NCDAAs relies on the BsrV enzyme, a periplasmic broad spectrum racemase...
September 2017: International Microbiology: the Official Journal of the Spanish Society for Microbiology
Masaki Osawa, Harold P Erickson
Bacterial cytokinesis begins with the assembly of FtsZ into a Z ring at the center of the cell. The Z-ring constriction in Gram-negative bacteria may occur in an environment where the periplasm and the cytoplasm are isoosmotic, but in Gram-positive bacteria the constriction may have to overcome a substantial turgor pressure. We address three potential sources of invagination force. (1) FtsZ itself may generate force by curved protofilaments bending the attached membrane. This is sufficient to constrict liposomes in vitro ...
2018: Frontiers in Microbiology
Wipa Suginta, Natchanok Sritho, Araya Ranok, David Bulmer, Yoshihito Kitaoku, Bert van den Berg, Tamo Fukamizo
Periplasmic solute-binding proteins (SBPs) in bacteria are involved in the active transport of nutrients into the cytoplasm. In marine bacteria of the genus Vibrio, a chitooligosaccharide-binding protein (CBP) is thought to be the major SBP controlling the rate of chitin uptake in these bacteria. However, the molecular mechanism of the CBP involvement in chitin metabolism has not been elucidated. Here, we report the structure and function of a recombinant chitooligosaccharide-binding protein from Vibrio harveyi , namely Vh CBP, expressed in Escherichia coli...
February 14, 2018: Journal of Biological Chemistry
Christopher D Radka, Lauren L Radford, Adriana V F Massicano, Lawrence J DeLucas, Suzanne E Lapi, Stephen G Aller
We demonstrate a scalable method for the separation of the bacterial periplasm from the cytoplasm. This method is used to purify periplasmic protein for the purpose of biophysical characterization, and measure substrate transfer between periplasmic and cytoplasmic compartments. By carefully limiting the time that the periplasm is separated from the cytoplasm, the experimenter can extract the protein of interest and assay each compartment individually for substrate without carry-over contamination between compartments...
February 1, 2018: Journal of Visualized Experiments: JoVE
Claudia A Studdert, Diego A Massazza
Chemoreceptors are dimeric proteins that contain a periplasmic or extracellular domain for ligand binding and an extremely well-conserved cytoplasmic domain for output response control. This latter domain consists in a long α-helical hairpin that forms a four-helix coiled-coil bundle in the dimer. Dimers associate into trimers of dimers in the crystal structure obtained for the cytoplasmic domain of the Escherichia coli serine chemoreceptor, Tsr. Further studies confirmed that this crystal structure reflects the basic unit within the in vivo organization of chemoreceptors...
2018: Methods in Molecular Biology
T L Leighton, M C Mok, M S Junop, P L Howell, L L Burrows
Pseudomonas aeruginosa uses long, thin fibres called type IV pili (T4P) for adherence to surfaces, biofilm formation, and twitching motility. A conserved subcomplex of PilMNOP is required for extension and retraction of T4P. To better understand its function, we attempted to co-crystallize the soluble periplasmic portions of PilNOP, using reductive surface methylation to promote crystal formation. Only PilOΔ109 crystallized; its structure was determined to 1.7 Å resolution using molecular replacement. This new structure revealed two novel features: a shorter N-terminal α1-helix followed by a longer unstructured loop, and a discontinuous β-strand in the second αββ motif, mirroring that in the first motif...
February 8, 2018: Scientific Reports
Marija Drikic, Jeroen De Buck
In health care, biosensors are envisioned as universal diagnostic devices with AAAA characteristics (i.e. available for anything, anywhere, anytime, to anyone). Despite numerous attempts to develop such a diagnostic device, none have managed to fulfill all 4 criteria and be commercialized. Glucometers, the most successful class of biosensor currently marketed monitor blood glucose concentrations. Their performance in clinical samples, including sensitivity and specificity, has been optimized and they are small and relatively inexpensive...
February 7, 2018: Biotechnology and Bioengineering
Abraham Rimon, Manish Dwivedi, Assaf Friedler, Etana Padan
Na+/H+ antiporters have a crucial role in pH- and Na+ homeostasis in cells. The crystal structure of NhaA, the main antiporter of Escherichia coli, has provided general insights into antiporter mechanisms and revealed a previously unknown structural fold, which has since been identified in several secondary active transporters. This unique structural fold is very delicately electrostatically balanced. Asp133 and Lys 300 have been ascribed essential roles in this balance, and, more generally, in the structure and function of the antiporter...
February 1, 2018: Journal of Molecular Biology
Zhilin Yang, Heejun Choi, James C Weisshaar
The permeabilization of model lipid bilayers by cationic peptides has been studied extensively over decades, with the bee-sting toxin melittin perhaps serving as the canonical example. However, the relevance of these studies to the permeabilization of real bacterial membranes by antimicrobial peptides remains uncertain. Here, we employ single-cell fluorescence microscopy in a detailed study of the interactions of melittin with the outer membrane (OM) and the cytoplasmic membrane (CM) of live Escherichia coli...
January 23, 2018: Biophysical Journal
M Hüttener, A Prieto, S Aznar, M Dietrich, S Paytubi, A Juárez
In this report we show that bacterial plasmids that harbor the Tn10 transposon (i.e., the IncHI1 plasmid R27) modify expression of different Salmonella regulons responding to the presence of tetracycline (Tc) in the medium. By using as a model the Tc-dependent upregulation of the ibpAB operon (which belongs to the heat shock regulon), we have identified Tn10- tetA (coding for a Tc efflux pump) and adjacent tetC sequences as required for ibpAB upregulation. Characterization of transcripts in the tetAC region showed that tetA transcription can continue into tetC sequences, generating a long 3'UTR sequence, which can protect transcripts from RNA processing, thus increasing expression of TetA protein...
February 2, 2018: Environmental Microbiology Reports
Sujittra Sujittra, Kanit Vichitphan, Jaehong Han, Sukanda Vichitphan, Jutaporn Swangkaew
For biotechnological production of high valued β-D-hexyl glucoside, catalytic properties of Hanseniaspora thailandica BC9 β-glucosidase purified from the periplasmic fraction was studied, and the transglycosylation activity for the production of β-D-hexyl glucoside was optimized. The constitutive BC9 β-glucosidase exhibited maximum specific activity at pH 6.0 and 40°C, and the activity of BC9 β-glucosidase was not significantly inhibited by various metal ions. BC9 β-glucosidase didn't show a significant activity of cellobiose hydrolysis, but the activity was rather enhanced in the presence of sucrose and medium chain alcohols...
February 1, 2018: Journal of Microbiology and Biotechnology
Philippe Leone, Jennifer Roche, Maxence S Vincent, Quang Hieu Tran, Aline Desmyter, Eric Cascales, Christine Kellenberger, Christian Cambillau, Alain Roussel
Type IX secretion system (T9SS), exclusively present in the Bacteroidetes phylum, has been studied mainly in Flavobacterium johnsoniae and Porphyromonas gingivalis. Among the 18 genes, essential for T9SS function, a group of four, porK-N (P. gingivalis) or gldK-N (F. johnsoniae) belongs to a co-transcribed operon that expresses the T9SS core membrane complex. The central component of this complex, PorM (or GldM), is anchored in the inner membrane by a trans-membrane helix and interacts through the outer membrane PorK-N complex...
January 30, 2018: Nature Communications
Anushila Chatterjee, Mark R O'Brian
Under iron limitation, bacteria scavenge ferric (Fe3+ ) iron bound to siderophores or other chelates from the environment to fulfill their nutritional requirement. In gram-negative bacteria, the siderophore uptake system prototype consists of an outer membrane transporter, a periplasmic binding protein, and a cytoplasmic membrane transporter, each specific for a single ferric siderophore or siderophore family. Here, we show that spontaneous single gain-of-function missense mutations in outer membrane transporter genes of Bradyrhizobium japonicum were sufficient to confer on cells the ability to use synthetic or natural iron siderophores, suggesting that selectivity is limited primarily to the outer membrane and can be readily modified...
January 30, 2018: Molecular Microbiology
J Wendler, D Ehmann, L Courth, B O Schroeder, N P Malek, J Wehkamp
The antimicrobial peptide human ß-defensin 1 (hBD1) is continuously produced by epithelial cells in many tissues. Compared to other defensins, hBD1 has only minor antibiotic activity in its native state. After reduction of its disulfide bridges, however, it becomes a potent antimicrobial agent against bacteria, while the oxidized native form (hBD1ox) shows specific activity against Gram-negative bacteria. We show that the killing mechanism of hBD1ox depends on aerobic growth conditions and bacterial enzymes...
January 29, 2018: Infection and Immunity
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