scholarly journals Rhamnolipid-Induced Removal of Lipopolysaccharide from Pseudomonas aeruginosa: Effect on Cell Surface Properties and Interaction with Hydrophobic Substrates

2000 ◽  
Vol 66 (8) ◽  
pp. 3262-3268 ◽  
Author(s):  
Ragheb A. Al-Tahhan ◽  
Todd R. Sandrin ◽  
Adria A. Bodour ◽  
Raina M. Maier
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Cell Surface ◽  
Structural Changes ◽  
Fatty Acid Content ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Cell Surface Hydrophobicity ◽  
Bacterial Cells ◽  
Low Levels ◽  
Rhamnolipid Biosurfactant

ABSTRACT Little is known about the interaction of biosurfactants with bacterial cells. Recent work in the area of biodegradation suggests that there are two mechanisms by which biosurfactants enhance the biodegradation of slightly soluble organic compounds. First, biosurfactants can solubilize hydrophobic compounds within micelle structures, effectively increasing the apparent aqueous solubility of the organic compound and its availability for uptake by a cell. Second, biosurfactants can cause the cell surface to become more hydrophobic, thereby increasing the association of the cell with the slightly soluble substrate. Since the second mechanism requires very low levels of added biosurfactant, it is the more intriguing of the two mechanisms from the perspective of enhancing the biodegradation process. This is because, in practical terms, addition of low levels of biosurfactants will be more cost-effective for bioremediation. To successfully optimize the use of biosurfactants in the bioremediation process, their effect on cell surfaces must be understood. We report here that rhamnolipid biosurfactant causes the cell surface ofPseudomonas spp. to become hydrophobic through release of lipopolysaccharide (LPS). In this study, two Pseudomonas aeruginosa strains were grown on glucose and hexadecane to investigate the chemical and structural changes that occur in the presence of a rhamnolipid biosurfactant. Results showed that rhamnolipids caused an overall loss in cellular fatty acid content. Loss of fatty acids was due to release of LPS from the outer membrane, as demonstrated by 2-keto-3-deoxyoctonic acid and sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis and further confirmed by scanning electron microscopy. The amount of LPS loss was found to be dependent on rhamnolipid concentration, but significant loss occurred even at concentrations less than the critical micelle concentration. We conclude that rhamnolipid-induced LPS release is the probable mechanism of enhanced cell surface hydrophobicity.

scholarly journals Variability in Pseudomonas aeruginosa Lipopolysaccharide Expression during Crude Oil Degradation

2002 ◽  
Vol 68 (10) ◽  
pp. 5096-5103 ◽  
Author(s):  
R. Sean Norman ◽  
Roberto Frontera-Suau ◽  
Pamela J. Morris
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Cell Surface ◽  
Crude Oil ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Cell Surface Hydrophobicity ◽  
Colony Morphology ◽  
Oil Degradation ◽  
O Antigen ◽  
Crude Oil Degradation

ABSTRACT Bacterial utilization of crude oil components, such as the n-alkanes, requires complex cell surface adaptation to allow adherence to oil. To better understand microbial cell surface adaptation to growth on crude oil, the cell surface characteristics of two Pseudomonas aeruginosa strains, U1 and U3, both isolated from the same crude oil-degrading microbial community enriched on Bonny Light crude oil (BLC), were compared. Analysis of growth rates demonstrated an increased lag time for U1 cells compared to U3 cells. Amendment with EDTA inhibited U1 and U3 growth and degradation of the n-alkane component of BLC, suggesting a link between cell surface structure and crude oil degradation. U1 cells demonstrated a smooth-to-rough colony morphology transition when grown on BLC, while U3 cells exhibited rough colony morphology at the outset. Combining high-resolution atomic force microscopy of the cell surface and sodium dodecyl sulfate-polyacrylamide gel electrophoresis of extracted lipopolysaccharides (LPS), we demonstrate that isolates grown on BLC have reduced O-antigen expression compared with that of glucose-grown cells. The loss of O-antigen resulted in shorter LPS molecules, increased cell surface hydrophobicity, and increased n-alkane degradation.

scholarly journals lfnA from Pseudomonas aeruginosa O12 and wbuX from Escherichia coli O145 Encode Membrane-Associated Proteins and Are Required for Expression of 2,6-Dideoxy-2-Acetamidino-l-Galactose in Lipopolysaccharide O Antigen

2007 ◽  
Vol 190 (5) ◽  
pp. 1671-1679 ◽  
Author(s):  
Jerry D. King ◽  
Erin F. Mulrooney ◽  
Evgeny Vinogradov ◽  
Bernd Kneidinger ◽  
Kristen Mead ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Pseudomonas Aeruginosa ◽  
Cell Surface ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Opportunistic Pathogen ◽  
Multidrug Resistant ◽  
Nuclear Magnetic Resonance Analysis ◽  
O Antigen ◽  
O Antigens

ABSTRACT The rare sugar 2,6-dideoxy-2-acetamidino-l-galactose (l-FucNAm) is found only in bacteria and is a component of cell surface glycans in a number of pathogenic species, including the O antigens of Pseudomonas aeruginosa serotype O12 and Escherichia coli O145. P. aeruginosa is an important opportunistic pathogen, and the O12 serotype is associated with multidrug-resistant epidemic outbreaks. O145 is one of the classic non-O157 serotypes associated with Shiga toxin-producing, enterohemorrhagic E. coli. The acetamidino (NAm) moiety of l-FucNAm is of interest, because at neutral pH it contributes a positive charge to the cell surface, and we aimed to characterize the biosynthesis of this functional group. The pathway is not known, but expression of NAm-modified sugars coincides with the presence of a pseA homologue in the relevant biosynthetic locus. PseA is a putative amidotransferase required for synthesis of a NAm-modified sugar in Campylobacter jejuni. In P. aeruginosa O12 and E. coli O145, the pseA homologues are lfnA and wbuX, respectively, and we hypothesized that these genes function in l-FucNAm biosynthesis. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis, Western blotting, and nuclear magnetic resonance analysis of the lfnA mutant O-antigen structure indicated that the mutant expresses 2,6-dideoxy-2-acetamido-l-galactose (l-FucNAc) in place of l-FucNAm. The mutation could be complemented by expression of either His6-tagged lfnA or wbuX in trans, confirming that these genes are functional homologues and that they are required for NAm moiety synthesis. Both proteins retained their activity when fused to a His6 tag and localized to the membrane fraction. These data will assist future biochemical investigation of this pathway.

Identification of pilin pools in the membranes of Pseudomonas aeruginosa

1982 ◽  
Vol 152 (2) ◽  
pp. 687-691
Author(s):  
T H Watts ◽  
E A Worobec ◽  
W Paranchych
Keyword(s):  
Staphylococcus Aureus ◽  
Pseudomonas Aeruginosa ◽  
Gel Electrophoresis ◽  
Type Strain ◽  
Wild Type Strain ◽  
Polyacrylamide Gel Electrophoresis ◽  
Protein A ◽  
Sodium Dodecyl ◽  
Wild Type ◽  
Outer Membranes

The proteins of purified inner and outer membranes obtained from Pseudomonas aeruginosa strains PAK and PAK/2Pfs were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis, transferred to nitrocellulose, and treated with antiserum raised against pure pili. Bound antipilus antibodies were visualized by reaction with 125I-labeled protein A from Staphylococcus aureus. The results showed that there are pools of pilin in both the inner and outer membranes of P. aeruginosa and that the pool size in the multipiliated strain is comparable with that of the wild-type strain.

Phospholipase C (heat-labile hemolysin) of Pseudomonas aeruginosa: purification and preliminary characterization

1982 ◽  
Vol 152 (1) ◽  
pp. 239-245
Author(s):  
R M Berka ◽  
M L Vasil
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Phospholipase C ◽  
Specific Activity ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Size Exclusion ◽  
Tetradecyltrimethylammonium Bromide ◽  
Heat Labile ◽  
Hydrophobic Moiety ◽  
Culture Supernatants

Phospholipase C (heat-labile hemolysin) was purified from Pseudomonas aeruginosa culture supernatants to near homogeneity by ammonium sulfate precipitation followed by a novel application of DEAE-Sephacel chromatography. Enzymatic activity remained associated with DEAE-Sephacel even in the presence of 1 M NaCl, but was eluted with a linear gradient of 0 to 5% tetradecyltrimethylammonium bromide. Elution from DEAE-Sephacel was also obtained with 2% lysophosphatidylcholine, and to a lesser extent with 2% phosphorylcholine, but not at all with choline. The enzyme was highly active toward phospholipids possessing substituted ammonium groups (e.g., phosphatidycholine, lysophosphatidylcholine, and sphingomyelin); however, it had little if any activity toward phospholipids lacking substituted ammonium groups (e.g., phosphatidylethanolamine, phosphatidylserine, and phosphaditylglycerol). Collectively, these data suggest that phospholipase C from P. aeruginosa exhibits high affinity for substituted ammonium groups, but requires an additional hydrophobic moiety for optimum binding. The specific activity of the purified enzyme preparation increased 1,900-fold compared with that of culture supernatants. The molecular weight of the phospholipase C was estimated to be 78,000 by both sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Sephacryl S-200 column chromatography and was 76,000 by high-performance size exclusion chromatography. The isoelectric point was 5.5. Amino acid analysis showed that phospholipase C was rich in glycine, serine, threonine, aspartyl, glutamyl, and aromatic amino acids, but was cystine free.

Changes in Cell Surface Properties of Shiga Toxigenic Escherichia coli by Quercus infectoria G. Olivier

2009 ◽  
Vol 72 (8) ◽  
pp. 1699-1704 ◽  
Author(s):  
SUPAYANG PIYAWAN VORAVUTHIKUNCHAI ◽  
SAKOL SUWALAK
Keyword(s):  
Escherichia Coli ◽  
Cell Surface ◽  
Surface Properties ◽  
Cell Surface Hydrophobicity ◽  
Ethanolic Extract ◽  
Bacterial Cells ◽  
Electron Microscopic ◽  
Cell Surface Properties ◽  
E Coli ◽  
Quercus Infectoria

The effects of Quercus infectoria (family Fagaceae) nutgalls on cell surface properties of Shiga toxigenic Escherichia coli (STEC) were investigated with an assay of microbial adhesion to hydrocarbon. The surface of bacterial cells treated with Q. infectoria exhibited a higher level of cell surface hydrophobicity (CSH) toward toluene than did the surface of untreated cells. With 50% ethanolic extract, the CSH of the three strains of STEC O157:H7 treated with 4× MIC of the extract resulted in moderate or strong hydrophobicity, whereas at 2× MIC and MIC, the CSH of only one strain of E. coli O157:H7 was significantly affected. The 95% ethanolic extract had a significant effect on CSH of all three strains at both 4× MIC and 2× MIC but not at the MIC. The effect on bacterial CSH was less pronounced with the other STEC strains. At 4× MIC, the 50% ethanolic extract increased the CSH of all non-O157 STEC strains significantly. At 2× MIC and 4× MIC, the 95% ethanolic extract affected the CSH of E. coli O26:H11 significantly but did not affect E. coli O111:NM or E. coli O22. Electron microscopic examination revealed the loss of pili in the treated cells. The ability of Q. infectoria extract to modify hydrophobic domains enables this extract to partition the lipids of the bacterial cell membrane, rendering the membrane more permeable and allowing leakage of ions and other cell contents, which leads to cell death. Further studies are required to evaluate the effects of Q. infectoria extract in food systems or in vivo and provide support for the use of this extract as a food additive for control of these STEC pathogens.

scholarly journals Effects of High Intensity Ultrasound on Physiochemical and Structural Properties of Goat Milk β-Lactoglobulin

Molecules ◽  
2020 ◽  
Vol 25 (16) ◽  
pp. 3637
Author(s):  
Xinhui Zhou ◽  
Cuina Wang ◽  
Xiaomeng Sun ◽  
Zixuan Zhao ◽  
Mingruo Guo
Keyword(s):  
Thermal Stability ◽  
Structural Properties ◽  
High Intensity ◽  
Structural Changes ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Goat Milk ◽  
High Intensity Ultrasound ◽  
Hydrophobicity Index ◽  
Β Lactoglobulin

This study aimed to compare the effects of high intensity ultrasound (HIU) applied at various amplitudes (20~40%) and for different durations (1~10 min) on the physiochemical and structural properties of goat milk β-lactoglobulin. No significant change was observed in the protein electrophoretic patterns by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Deconvolution and second derivative of the Fourier transform infrared spectra (FTIR) showed that the percentage of β-sheet of goat milk β-lactoglobulin was significantly decreased while those of α-helix and random coils increased after HIU treatment The surface hydrophobicity index and intrinsic fluorescence intensity of samples was enhanced and increased with increasing HIU amplitude or time. Differential scanning calorimetry (DSC) results exhibited that HIU treatments improved the thermal stability of goat milk β-lactoglobulin. Transmission electron microscopy (TEM) of samples showed that the goat milk β-lactoglobulin microstructure had changed and it contained larger aggregates when compared with the untreated goat milk β-lactoglobulin sample. Data suggested that HIU treatments resulted in secondary and tertiary structural changes of goat milk β-lactoglobulin and improved its thermal stability.

Rapid detection and initial characterization of genetic variants of human serum albumin

1991 ◽  
Vol 37 (7) ◽  
pp. 1221-1224 ◽  
Author(s):  
J Merle Sheat ◽  
Robert J Peach ◽  
Peter M George
Keyword(s):  
Human Serum Albumin ◽  
Serum Albumin ◽  
Human Serum ◽  
Genetic Variants ◽  
Gel Electrophoresis ◽  
Structural Changes ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl

Abstract We have studied the detection and classification of genetic variants of human serum albumin by electrophoresis. Samples from 10 patients who were heterozygous for eight different albumin variants were studied by two methods. In agarose gel electrophoresis, each of these variants has an abnormal mobility and can be classified on the basis that structural changes at the N-terminus abolish 63Ni binding. In sodium dodecyl sulfate-polyacrylamide gel electrophoresis of whole serum, glycosylated variants are easily detected because of their greater apparent molecular mass.

scholarly journals Cell-surface radioiodination with the sparingly soluble catalyst Iodogen. Difference between the dividing and non-dividing populations of rodent thymocytes

1981 ◽  
Vol 194 (1) ◽  
pp. 351-355 ◽  
Author(s):  
J G Salisbury ◽  
J M Graham
Keyword(s):  
Cell Surface ◽  
Sodium Dodecyl Sulphate ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Polyacrylamide Gel ◽  
Surface Proteins ◽  
New Method

The surface proteins of dividing and non-dividing subpopulations of rat and mouse thymocytes have been labelled by using a new method of radioiodination. Sodium dodecyl sulphate/polyacrylamide-gel electrophoresis and autoradiography of the labelled proteins shows distinct differences in labelling between the mouse and rat cells and also, in the case of the rat, between the dividing and non-dividing populations.

scholarly journals Motility and the Polar Flagellum Are Required for Aeromonas caviae Adherence to HEp-2 Cells

2001 ◽  
Vol 69 (7) ◽  
pp. 4257-4267 ◽  
Author(s):  
Ali A. Rabaan ◽  
Ioannis Gryllos ◽  
Juan M. Tomás ◽  
Jonathan G. Shaw
Keyword(s):  
Epithelial Cells ◽  
Molecular Mass ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Kanamycin Resistance ◽  
Polar Flagellum ◽  
Bacterial Cells ◽  
Aeromonas Caviae ◽  
Human Epithelial Cells

ABSTRACT Aeromonas caviae is increasingly being recognized as a cause of gastroenteritis, especially among the young. The adherence of aeromonads to human epithelial cells in vitro has been correlated with enteropathogenicity, but the mechanism is far from well understood. Initial investigations demonstrated that adherence of A. caviae to HEp-2 cells was significantly reduced by either pretreating bacterial cells with an antipolar flagellin antibody or by pretreating HEp-2 cells with partially purified flagella. To precisely define the role of the polar flagellum in aeromonad adherence, we isolated the A. caviae polar flagellin locus and identified five polar flagellar genes, in the order flaA, flaB, flaG, flaH, and flaJ. Each gene was inactivated using a kanamycin resistance cartridge that ensures the transcription of downstream genes, and the resulting mutants were tested for motility, flagellin expression, and adherence to HEp-2 cells. N-terminal amino acid sequencing, mutant analysis, and Western blotting demonstrated that A. caviae has a complex flagellum filament composed of two flagellin subunits encoded by flaAand flaB. The predicted molecular mass of both flagellins was ∼31,700 Da; however, their molecular mass estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis was ∼35,500 Da. This aberrant migration was thought to be due to their glycosylation, since the proteins were reactive in glycosyl group detection assays. Single mutations in either flaA orflaB did not result in loss of flagella but did result in decreased motility and adherence by approximately 50%. Mutation offlaH, flaJ, or both flagellin genes resulted in the complete loss of motility, flagellin expression, and adherence. However, mutation of flaG did not affect motility but did significantly reduce the level of adherence. Centrifugation of the flagellate mutants (flaA, flaB, and flaG) onto the cell monolayers did not increase adherence, whereas centrifugation of the aflagellate mutants (flaH, flaJ, and flaA flaB) increased adherence slightly. We conclude that maximum adherence of A. caviae to human epithelial cells in vitro requires motility and optimal flagellar function.

Cell surface characteristics of Mortierella species and their interaction with a mycoparasite

1984 ◽  
Vol 30 (3) ◽  
pp. 290-298 ◽  
Author(s):  
M. S. Manocha
Keyword(s):  
Cell Wall ◽  
Cell Surface ◽  
Periodic Acid ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Surface Characteristics ◽  
Protein Patterns ◽  
Periodic Acid Schiff ◽  
The Difference ◽  
Chitin And Chitosan

Cell surface characteristics of three Mortierella species differing in their response to a mycoparasite, Piptocephalis virginiana, were examined. Their cell wall composition was typical of mucoraceous fungi with chitin and chitosan as major polysaccharides. Electron microscopy revealed that the mycoparasite penetrated and formed haustoria in the hyphae of susceptible hosts, M. pusilla and M. isabellina. The failure of the parasite to establish contact and penetrate a hypha of the nonhost, M. candelabrum, was not due to cell wall thickness, rigidity, or chitin contents. Markedly different protein patterns obtained from crude alkali extracts of host and nonhost cell walls by sodium dodecyl sulfate – polyacrylamide gel electrophoresis might explain the difference in host and nonhost response to the mycoparasite. Whereas most of the bands differed only in intensity after staining with either Coomassie blue or periodic acid – Schiff reagent, there were two distinct bands of glycoproteins (76 000 and 74 000) observed in the host species which were absent in the nonhost species.

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