scholarly journals Biosynthesis and turnover of outer-membrane proteins in Escherichia coli ML308-225

1979 ◽  
Vol 182 (2) ◽  
pp. 407-412 ◽  
Author(s):  
R J Allen ◽  
G K Scott
Keyword(s):  
Escherichia Coli ◽  
Protein Synthesis ◽  
Outer Membrane ◽  
Bacterial Growth ◽  
Growth Phase ◽  
Exponential Growth ◽  
Outer Membrane Proteins ◽  
Exponential Growth Phase ◽  
Outer Membranes ◽  
Dilution Experiments

Isolated outer membranes and outer-membrane extracts from Escherichia coli ML308-225 in the early-exponential growth phase contain more protein than do corresponding preparations from late-exponential- or stationary-phase bacteria. Isotope-dilution experiments show that this is due to a loss of protein from the membrane during the exponential growth phase. Inhibition of bacterial growth and protein synthesis stabilizes the outer-membrane-protein concentration. Protein synthesis in the absence of bacterial growth results in higher concentrations of protein in the outer membrane.

scholarly journals Expression and Porin Activity of P28 and OMP-1F during Intracellular Ehrlichia chaffeensis Development

2008 ◽  
Vol 190 (10) ◽  
pp. 3597-3605 ◽  
Author(s):  
Yumi Kumagai ◽  
Haibin Huang ◽  
Yasuko Rikihisa
Keyword(s):  
Nutrient Uptake ◽  
Outer Membrane ◽  
Growth Phase ◽  
Exponential Growth ◽  
Outer Membrane Proteins ◽  
Exponential Growth Phase ◽  
Ehrlichia Chaffeensis ◽  
Negative Bacterium ◽  
Gram Negative ◽  
Pore Sizes

ABSTRACT Ehrlichia chaffeensis, an obligatory intracellular gram-negative bacterium, must take up various nutrients and metabolic compounds because it lacks many genes involved in metabolism. Nutrient uptake by a gram-negative bacterium occurs primarily through pores or channels in the bacterial outer membrane. Here we demonstrate that isolated E. chaffeensis outer membranes have porin activities, as determined by a proteoliposome swelling assay. The activity was partially blocked by an antibody that recognizes the two most abundant outer membrane proteins, P28/OMP-19 and OMP-1F/OMP-18. Both proteins were predicted to have structural features characteristic of porins, including 12 transmembrane segments comprised of amphipathic and antiparallel β-strands. The sodium dodecyl sulfate stability of the two proteins was consistent with a β-barrel structure. Isolated native P28 and OMP-1F exhibited porin activities, with pore sizes similar to and larger than, respectively, that of OprF, which is the porin with the largest pore size known to date. E. chaffeensis experiences temperature changes during transmission by ticks. During the intracellular development of E. chaffeensis, both P28 and OMP-1F were expressed mostly in the mid-exponential growth phase at 37°C and the late-exponential growth phase at 28°C. The porin activity of proteoliposomes reconstituted with proteins from the outer membrane fractions derived from bacteria in the mid- and late-exponential growth phases at 28°C and 37°C correlated with the expression levels of P28 and OMP-1F. These results imply that P28 and OMP-1F function as porins with large pore sizes, suggesting that the differential expression of these two proteins might regulate nutrient uptake during intracellular E. chaffeensis development at both temperatures.

scholarly journals Azithromycin Exhibits Bactericidal Effects on Pseudomonas aeruginosa through Interaction with the Outer Membrane

2005 ◽  
Vol 49 (4) ◽  
pp. 1377-1380 ◽  
Author(s):  
Yoshifumi Imamura ◽  
Yasuhito Higashiyama ◽  
Kazunori Tomono ◽  
Koichi Izumikawa ◽  
Katsunori Yanagihara ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Outer Membrane ◽  
Growth Phase ◽  
Exponential Growth ◽  
Divalent Cations ◽  
Stationary Growth Phase ◽  
Exponential Growth Phase ◽  
Stationary Growth ◽  
Outer Membranes ◽  
Bactericidal Effects

ABSTRACT The aim of the present study was to elucidate the effect of the macrolide antibiotic azithromycin on Pseudomonas aeruginosa. We studied the susceptibility to azithromycin in P. aeruginosa PAO1 using a killing assay. PAO1 cells at the exponential growth phase were resistant to azithromycin. In contrast, PAO1 cells at the stationary growth phase were sensitive to azithromycin. The divalent cations Mg2+ and Ca2+ inhibited this activity, suggesting that the action of azithromycin is mediated by interaction with the outer membranes of the cells, since the divalent cations exist between adjacent lipopolysaccharides (LPSs) and stabilize the outer membrane. The divalent cation chelator EDTA behaved in a manner resembling that of azithromycin; EDTA killed more PAO1 in the stationary growth phase than in the exponential growth phase. A 1-N-phenylnaphthylamine assay showed that azithromycin interacted with the outer membrane of P. aeruginosa PAO1 and increased its permeability while Mg2+ and Ca2+ antagonized this action. Our results indicate that azithromycin directly interacts with the outer membrane of P. aeruginosa PAO1 by displacement of divalent cations from their binding sites on LPS. This action explains, at least in part, the effectiveness of sub-MICs of macrolide antibiotics in pseudomonal chronic airway infection.

scholarly journals The Putrescine Importer PuuP of Escherichia coli K-12

2009 ◽  
Vol 191 (8) ◽  
pp. 2776-2782 ◽  
Author(s):  
Shin Kurihara ◽  
Yuichi Tsuboi ◽  
Shinpei Oda ◽  
Hyeon Guk Kim ◽  
Hidehiko Kumagai ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Gene Cluster ◽  
Growth Phase ◽  
Exponential Growth ◽  
Minimal Medium ◽  
Exponential Growth Phase ◽  
E Coli ◽  
Genes Encoding ◽  
K 12 ◽  
Utilization Pathway

ABSTRACT The Puu pathway is a putrescine utilization pathway involving gamma-glutamyl intermediates. The genes encoding the enzymes of the Puu pathway form a gene cluster, the puu gene cluster, and puuP is one of the genes in this cluster. In Escherichia coli, three putrescine importers, PotFGHI, PotABCD, and PotE, were discovered in the 1990s and have been studied; however, PuuP had not been discovered previously. This paper shows that PuuP is a novel putrescine importer whose kinetic parameters are equivalent to those of the polyamine importers discovered previously. A puuP + strain absorbed up to 5 mM putrescine from the medium, but a ΔpuuP strain did not. E. coli strain MA261 has been used in previous studies of polyamine transporters, but PuuP had not been identified previously. It was revealed that the puuP gene of MA261 was inactivated by a point mutation. When E. coli was grown on minimal medium supplemented with putrescine as the sole carbon or nitrogen source, only PuuP among the polyamine importers was required. puuP was expressed strongly when putrescine was added to the medium or when the puuR gene, which encodes a putative repressor, was deleted. When E. coli was grown in M9-tryptone medium, PuuP was expressed mainly in the exponential growth phase, and PotFGHI was expressed independently of the growth phase.

Induction of autolysis in Bacillus subtilis by ochratoxin A

1978 ◽  
Vol 24 (5) ◽  
pp. 563-568 ◽  
Author(s):  
U. Singer ◽  
R. Röschenthaler
Keyword(s):  
Cell Wall ◽  
Protein Synthesis ◽  
Bacillus Subtilis ◽  
Ochratoxin A ◽  
Optical Density ◽  
Growth Phase ◽  
Exponential Growth ◽  
Rna Synthesis ◽  
Exponential Growth Phase ◽  
Cell Wall Synthesis

Ochratoxin A (OTA) added during the exponential growth phase at a concentration higher than 12 μg/ml caused autolysis of Bacillus subtilis. Optical density of cultures decreased, and at higher concentrations the cultures became sterile. Optimum OTA-induced lysis was about pH 5. At concentrations below 10 μg/ml, protein synthesis was inhibited more strongly than RNA synthesis. Cell wall synthesis was also strongly inhibited. A fraction extracted from the lysates had the property of a lysis inhibitor. The relevance of this fraction in respect to autolysis is discussed.

scholarly journals Adhesion of Type 1-Fimbriated Escherichia coli to Abiotic Surfaces Leads to Altered Composition of Outer Membrane Proteins

2001 ◽  
Vol 183 (8) ◽  
pp. 2445-2453 ◽  
Author(s):  
Karen Otto ◽  
Joakim Norbeck ◽  
Thomas Larsson ◽  
Karl-Anders Karlsson ◽  
Malte Hermansson
Keyword(s):  
Escherichia Coli ◽  
Protein Synthesis ◽  
Membrane Proteins ◽  
Outer Membrane ◽  
Cell Envelope ◽  
Outer Membrane Proteins ◽  
Polyacrylamide Gel Electrophoresis ◽  
Surface Characteristics ◽  
Abiotic Surfaces

ABSTRACT Phenotypic differences between planktonic bacteria and those attached to abiotic surfaces exist, but the mechanisms involved in the adhesion response of bacteria are not well understood. By the use of two-dimensional (2D) polyacrylamide gel electrophoresis, we have demonstrated that attachment of Escherichia coli to abiotic surfaces leads to alteration in the composition of outer membrane proteins. A major decrease in the abundance of resolved proteins was observed during adhesion of type 1-fimbriated E. colistrains, which was at least partly caused by proteolysis. Moreover, a study of fimbriated and nonfimbriated mutants revealed that these changes were due mainly to type 1 fimbria-mediated surface contact and that only a few changes occurred in the outer membranes of nonfimbriated mutant strains. Protein synthesis and proteolytic degradation were involved to different extents in adhesion of fimbriated and nonfimbriated cells. While protein synthesis appeared to affect adhesion of only the nonfimbriated strain, proteolytic activity mostly seemed to contribute to adhesion of the fimbriated strain. Using matrix-assisted laser desorption ionization–time of flight mass spectrometry, six of the proteins resolved by 2D analysis were identified as BtuB, EF-Tu, OmpA, OmpX, Slp, and TolC. While the first two proteins were unaffected by adhesion, the levels of the last four were moderately to strongly reduced. Based on the present results, it may be suggested that physical interactions between type 1 fimbriae and the surface are part of a surface-sensing mechanism in which protein turnover may contribute to the observed change in composition of outer membrane proteins. This change alters the surface characteristics of the cell envelope and may thus influence adhesion.

scholarly journals TonB Interacts with Nonreceptor Proteins in the Outer Membrane of Escherichia coli

2002 ◽  
Vol 184 (6) ◽  
pp. 1640-1648 ◽  
Author(s):  
Penelope I. Higgs ◽  
Tracy E. Letain ◽  
Kelley K. Merriam ◽  
Neal S. Burke ◽  
HaJeung Park ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Outer Membrane ◽  
Cytoplasmic Membrane ◽  
Outer Membrane Proteins ◽  
Membrane Receptors ◽  
Strong Interactions ◽  
Cross Linking ◽  
Outer Membrane Receptor ◽  
Outer Membranes

ABSTRACT The Escherichia coli TonB protein serves to couple the cytoplasmic membrane proton motive force to active transport of iron-siderophore complexes and vitamin B12 across the outer membrane. Consistent with this role, TonB has been demonstrated to participate in strong interactions with both the cytoplasmic and outer membranes. The cytoplasmic membrane determinants for that interaction have been previously characterized in some detail. Here we begin to examine the nature of TonB interactions with the outer membrane. Although the presence of the siderophore enterochelin (also known as enterobactin) greatly enhanced detectable cross-linking between TonB and the outer membrane receptor, FepA, the absence of enterochelin did not prevent the localization of TonB to the outer membrane. Furthermore, the absence of FepA or indeed of all the iron-responsive outer membrane receptors did not alter this association of TonB with the outer membrane. This suggested that TonB interactions with the outer membrane were not limited to the TonB-dependent outer membrane receptors. Hydrolysis of the murein layer with lysozyme did not alter the distribution of TonB, suggesting that peptidoglycan was not responsible for the outer membrane association of TonB. Conversely, the interaction of TonB with the outer membrane was disrupted by the addition of 4 M NaCl, suggesting that these interactions were proteinaceous. Subsequently, two additional contacts of TonB with the outer membrane proteins Lpp and, putatively, OmpA were identified by in vivo cross-linking. These contacts corresponded to the 43-kDa and part of the 77-kDa TonB-specific complexes described previously. Surprisingly, mutations in these proteins individually did not appear to affect TonB phenotypes. These results suggest that there may be multiple redundant sites where TonB can interact with the outer membrane prior to transducing energy to the outer membrane receptors.

A particulate chitin synthase from Aspergillus flavus Link: the properties, location, and levels of activity in mycelium and regenerating protoplast preparations

1976 ◽  
Vol 22 (7) ◽  
pp. 915-921 ◽  
Author(s):  
P. M. Moore ◽  
J. F. Peberdy
Keyword(s):  
Protein Synthesis ◽  
Carbon Source ◽  
Aspergillus Flavus ◽  
Growth Phase ◽  
Exponential Growth ◽  
Incubation Temperature ◽  
Specific Activity ◽  
Chitin Synthase ◽  
Exponential Growth Phase ◽  
Net Protein

Chitin synthase (EC 2.4.1.16) has been characterized in Aspergillus flavus. A Km value of 2.5 mM was obtained for the substrate UDPGlcNAc. The enzyme had a requirement for GlcNAc, and Mg2+ and activity was increased in the presence of soluble chitodextrins F1 and F2. The optimum activity was obtained using Tris–HCl buffer, pH 7.5, with a secondary peak at pH 6.2 and an incubation temperature of 29.5 °C.Distribution patterns of chitin synthase in protoplasts and mycelial material were very similar. The highest specific activity was found in a 200 000 × g fraction.Enzyme levels in growing mycelium increased during the exponential growth phase after which they declined. Activity also increased during the early stages of regeneration of both conidial and mycelial protoplasts, despite an initial lack in net protein synthesis. Chitin synthase levels were also dependent upon the carbon source available during regeneration.

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