Hot Sodium Dodecyl Research Articles

Covalent Biofunctionalization of the Inner Surfaces of a Hollow-Fiber Capillary Bundle Using Packed-Bed Plasma Ion Implantation.

Hollow-fiber capillary bundles are widely used in the production of medical devices for blood oxygenation and purification purposes such as in cardiopulmonary bypass, hemodialysis, and hemofiltration, but the blood interfacing inner surfaces of these capillaries provide poor hemocompatibility. Here, we present a novel method of packed-bed plasma ion implantation (PBPII) for the modification of the inner surfaces of polymeric hollow-fiber bundles enclosed in a cassette. The method is simple and can be performed on an intact hollow-fiber bundle cassette by the placement of a hollow cylindrical electrode, connected to a negative high-voltage pulse generator, around the cassette. The method does not require the insertion of electrodes inside the capillaries or the cassette. Nitrogen gas is fed into the capillaries inside the cassette by connecting the inlet of the cassette to a gas source. Upon the application of negative high-voltage bias pulses to the electrode, plasma is ignited inside the cassette, achieving the surface modification of both the internal and external surfaces of the capillaries. Fourier transform infrared-attenuated total reflectance spectroscopy of the PBPII-treated capillaries revealed the formation of aromatic C═C bonds, indicating the progressive carbonization of the capillary surfaces. The PBPII treatment was found to be uniform along the capillaries and independent of the radial position in the cassette. Atomic force microscopy of cross sections through the capillaries revealed that the increased stiffness associated with the carbonized layer on the inner surface of the PBPII-treated capillary has a depth (∼40 nm) consistent with that expected for ions accelerated by the applied bias voltage. The modified internal surfaces of the capillary bundle showed a greatly increased wettability and could be biofunctionalized by covalently immobilizing protein directly from the buffer solution. The direct, reagent-free protein immobilization was demonstrated using tropoelastin as an example protein. Covalent binding of the protein was confirmed by its resistance to removal by hot sodium dodecyl sulfate detergent washing, which is known to disrupt physical binding.

Plasma activated coatings with dual action against fungi and bacteria

In the oral cavity, dental implants are exposed to an environment rich in various microbes that can produce infectious biofilms on the implant surface. Here we report the development of two distinct antimicrobial coatings that prevent biofilm formation by fungi or bacteria. The antimicrobial peptides Mel4 and caspofungin were immobilized on titanium surfaces through reactions with radicals embedded within a mechanically robust, ion-assisted plasma polymerized (PP) film. The immobilization does not require additional chemical reagents and is achieved by simply incubating the surfaces at room temperature in a buffer solution containing the antimicrobial agent. The antibiotic-functionalized surfaces were rigorously washed with hot sodium dodecyl sulphate (SDS) to remove physisorbed molecules, and analyzed by time of flight secondary ion mass spectrometry (ToF-SIMS), which revealed characteristic fragments of the peptides and provided strong evidence for the covalent nature of the binding between the molecules and the PP coating. Both Candida albicans and Staphylococcus aureus pathogens were significantly inhibited in their ability to colonize the surfaces and form biofilms. Our findings suggest that antimicrobial surfaces fabricated using ion-assisted plasma polymerization have great potential for coatings on biomedical devices where activity against fungal and bacterial pathogens is required.

Characterization of Lipids and Proteins Associated to the Cell Wall of the Acapsular Mutant Cryptococcus neoformans Cap 67.

Cryptococcus neoformans is an opportunistic human pathogen that causes life-threatening meningitis. In this fungus, the cell wall is exceptionally not the outermost structure due to the presence of a surrounding polysaccharide capsule, which has been highly studied. Considering that there is little information about C. neoformans cell wall composition, we aimed at describing proteins and lipids extractable from this organelle, using as model the acapsular mutant C. neoformans cap 67. Purified cell wall preparations were extracted with either chloroform/methanol or hot sodium dodecyl sulfate. Total lipids fractionated in silica gel 60 were analyzed by electrospray ionization tandem mass spectrometry (ESI-MS/MS), while trypsin digested proteins were analyzed by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). We detected 25 phospholipid species among phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, phosphatidylinositol, and phosphatidic acid. Two glycolipid species were identified as monohexosyl ceramides. We identified 192 noncovalently linked proteins belonging to different metabolic processes. Most proteins were classified as secretory, mainly via nonclassical mechanisms, suggesting a role for extracellular vesicles (EV) in transwall transportation. In concert with that, orthologs from 86% of these proteins have previously been reported both in fungal cell wall and/or in EV. The possible role of the presently described structures in fungal-host relationship is discussed.

Isolation and Physical Characterization of Hydrophobin-Like Proteins (HLP) from Aerial Conidia of <i>Metarhizium Anisopliae</i> Var. <i>Anisopliae</i>

The aim of this study was to isolate and physically characterized Hyrophobin-Like Proteins (HLP) from the aerial conidia of two local isolates of Metarhizium anisopliae var. anisopliae, namely, TA and LR2. The protein samples were isolated based on their insolubility in hot Sodium Dodecyl Sulfate (SDS). The SDS-insoluble proteins were then purified in cold 98% formic acid and performic acid. The molecular weight of the Formic Acid Extracted (FAE) proteins fell in the range of 13-17 kDa with the contents of 3.80 and 3.56 mg mL-1 for TA and LR2, respectively. Due to the stringent protocol of isolation and purification, the FAE proteins can be verified as Hydrophobin-Like Proteins (HLP). Both the HLP samples isolated from M. anisopliae isolates TA and LR2 have low contact angles of 57.06±0.38 and 58.43±0.25°, respectively. The HLP also revealed good emulsification effect in the oil-water phase. Conidia showed good dispersion in aqueous solution with the application of HLP. Paper pre-coated with HLP resisted wetting by water for up to 439 sec. These unique properties of HLP from local isolates of M. anisopliae var. anisopliae are of great potential to be used in wide range of industrial applications.

Toluene gas phase biofiltration by Paecilomyces lilacinus and isolation and identification of a hydrophobin protein produced thereof

Paecilomyces lilacinus consumed toluene as the sole carbon source in a gas-phase biofilter packed with perlite obtaining an average elimination capacity of 50 g m(-3) h(-1), a removal efficiency of 53%, and a final biomass of 31.6 mg biomass g dry support(-1). Hydrophobin proteins from the mycelium produced in the biofilter were purified by formic acid extraction and precipitated by electrobubbling, and the molecular weight was found to be 10.6 +/- 0.3 kDa. The peptide mass fingerprinting analysis of the purified hydrophobin by matrix-assisted laser desorption/ionization time-of-flight resulted in the identification of two peptides that presented high homology with sequences of class I hydrophobin proteins from other ascomycetous fungi when compared against the National Center for Biotechnology Information database. The yield of hydrophobin (PLHYD) from P. lilacinus was 1.1 mg PLHYD g biomass(-1). These proteins modified the hydrophobicity of Teflon by lowering the contact angle from 130.1 (+/-2) degrees to 57.0 (+/-5) degrees supporting hot sodium dodecyl sulfate washing. This work is the first report about biodegradation of toluene by the nematophagous fungus P. lilacinus in a gas-phase biofilter and the identification of its hydrophobin protein.

Isolation and Quantitative Estimation of Diesel Exhaust and Carbon Black Particles Ingested by Lung Epithelial Cells and Alveolar Macrophages in Vitro

A new procedure for isolating and estimating ingested carbonaceous diesel exhaust particles (DEP) or carbon black (CB) particles by lung epithelial cells and macrophages is described. Cells were incubated with DEP or CB to examine cell-particle interaction and ingestion. After various incubation periods, the cells were separated from free extracellular DEP or CB particles by Ficoll density gradient centrifugation and dissolved in hot sodium dodecyl sulfate detergent. Insoluble DEP or CB residues were isolated by high-speed centrifugation, and the elemental carbon (EC) concentrations in the pellets were estimated by a thermal-optical-transmittance method (i.e., carbon analysis). From the EC concentration, the amount of ingested DEP or CB could be calculated. The described technique allowed the determination of the kinetics and dose dependence of DEP uptake by LA4 lung epithelial cells and MHS alveolar macrophages. Both cell types ingested DEP to a similar degree; however, the MHS macrophages took up significantly more CB than the epithelial cells. Cytochalasin D, an agent that blocks actin polymerization in the cells, inhibited approximately 80% of DEP uptake by both cell types, indicating that the process was actin-dependent in a manner similar to phagocytosis. This technique can be applied to examine the interactions between cells and particles containing EC and to study the modulation of particle uptake in diseased tissue.

Extraction of bulk DNA from Thar Desert soils for optimization of PCR-DGGE based microbial community analysis

A reliable method for characterizing microbial communities on the basis of their differences in the 16S ribosomal RNA (rRNA) gene sequences in the hot arid zone sandy soils has been optimized. A desert plant (Calligonum polygonoides) was chosen to provide the rhizospheric soil samples, collected from three different agro-ecological locations. Total community DNA was efficiently extracted at small-scale level using direct lysis with hot sodium dodecyl sulphate (SDS), glass bead beating and finally subjecting the sandy soil to liquid nitrogen freeze-thaw cycles. To amplify V3 region of bacterial 16S rRNA gene, universal conserved primers were used. Second round polymerase chain reaction (PCR) was attempted to increase product concentration and to minimize the effect of inhibitory substances. To enhance the detection sensitivity of the denaturing gradient gel electrophoresis (DGGE), the effect of change in template DNA concentration was studied. The separation of bands were greatly enhanced in the fingerprints obtained after the second round of PCR representing low abundant species which were not differentiated at single optimized concentration of DNA

Investigation ofFasciola hepatica sample preparation for two-dimensional electrophoresis

This paper investigates the preparation of Fasciola hepatica samples for two-dimensional electrophoresis (2-DE). Whole samples were prepared by both hot sodium dodecyl sulfate (SDS) solubilisation and precipitation using trichloroacetic acid (TCA) to remove nonprotein contaminants and to inactivate endogenous proteases. Sample preparation had a marked influence on the 2-DE gel profile. TCA precipitation resulted in no measurable improvement in the profile observed, compared to the untreated control. Solubilisation of sample with hot SDS increased the number of protein spots, as did TCA precipitation with the addition of phosphotungstic acid. The preparation of excretory-secretory (ES) products poses problems due to both high salt concentrations and low protein concentration. All precipitation methods used to overcome this gave similar profiles, except acetone alone, which caused depletion of the larger proteins. TCA in acetone gave the best result, similar to that obtained by centrifugal filtration of the sample. Overcrowding of spots in some regions of the 2-DE gel occurred in the whole Fasciola hepatica sample. This problem was alleviated by differential solubilisation, which also resulted in the enrichment of some proteins.

Deletion of new covalently linked cell wall glycoproteins alters the electrophoretic mobility of phosphorylated wall components of Saccharomyces cerevisiae.

The incorporation of radioactive orthophosphate into the cell walls of Saccharomyces cerevisiae was studied. 33P-labeled cell walls were extensively extracted with hot sodium dodecyl sulfate (SDS). Of the remaining insoluble radioactivity more than 90% could be released by laminarinase. This radioactive material stayed in the stacking gel during SDS-polyacrylamide gel electrophoresis but entered the separating gel upon treatment with N-glycosidase F, indicating that phosphate was linked directly or indirectly to N-mannosylated glycoproteins. The phosphate was bound to covalently linked cell wall proteins as mannose-6-phosphate, the same type of linkage shown previously for soluble mannoproteins (L. Ballou, L. M. Hernandez, E. Alvarado, and C. E. Ballou, Proc. Natl. Acad. Sci. USA 87:3368-3372, 1990). From the phosphate-labeled glycoprotein fraction released by laminarinase, three cell wall mannoproteins, Ccw12p, Ccw13p and Ccw14p, were isolated and identified by N-terminal sequencing. For Ccw13p (encoded by DAN1 [also called TIR3]) and Ccw12p the association with the cell wall has not been described before; Ccw14p is identical with cell wall protein Icwp (I. Moukadiri, J. Armero, A. Abad, R. Sentandreu, and J. Zueco, J. Bacteriol. 179:2154-2162, 1997). In ccw12, ccw13, or ccw14 single or double mutants neither the amount of radioactive phosphate incorporated into cell wall proteins nor its position in the stacking gel was changed. However, the triple mutant brought about a shift of the 33P-labeled glycoprotein components from the stacking gel into the separating gel. The disruption of CCW12 results in a pronounced sensitivity of the cells to calcofluor white and Congo red. In addition, the ccw12 mutant shows a decrease in mating efficiency and a defect in agglutination.

Reaggregation and binding of cell wall proteins from Candida albicans to structural polysaccharides

Urea or hot sodium dodecyl sulphate extracted a significant amount of the same proteins from the matrix of the cell wall of the yeast form and mycelial cells of Candida albicans. Gel filtration analysis of the urea-extracted proteins revealed that they occurred in the form of large complexes which were unaffected by up to 8 M urea. Among them, proteins en route to becoming covalently associated within the wall scaffold were identified by their reaction with specific antibodies. When urea was removed by dialysis, some of these proteins specifically reassociated into large aggregates which bound strongly with ConA, whereas others remained soluble in smaller associated products. The ability of some of these proteins to bind to the insoluble wall polysaccharides was also assessed. No self-assembling proteins were able to bind to glucans and/or chitin. Specificity of the binding to polysaccharides made of β-bound glucosyl or N-acetylglucosaminyl residues was determined by the competitive effect of several disaccharides. Whereas laminaribiose and diacetylchitobiose were strong inhibitors of protein binding to both glucan and chitin, lactose, maltose and sucrose were ineffective.

Lectin-oral streptococci interactions.

Lectins of various specificities were examined for interaction with strains of oral streptococci of various species. The lectins were capable of binding galactose, N-acetylgalactosamine, glucose, N-acetylglucosamine, mannose, fucose and sialic acid. Lectin reactivities were strain-dependent in that some members of a species, but not others, could be aggregated by certain lectins. Proteolysis and extraction with hot water, guanidine hydrochloride and sodium dodecyl sulphate tended to increase the reactivity of the streptococci with lectins but did not change the recognition patterns of the bacteria. Sonication, in contrast, tended to reduce the ability of streptococci to be agglutinated by lectins. Furthermore, lectin reactivities were dependent on the growth conditions, as evidenced by changes in lectin titres following streptococcal growth in sub-inhibitory concentrations of citrate, fluoride or antibiotics. It is likely that lectins could be useful tools for epidemiological studies and to probe strain-dependent and growth-dependent surface characteristics of viridans streptococci.

Role of the C terminus in antigen P1 surface localization in Streptococcus mutans and two related cocci.

The C terminus of the major surface protein P1 from Streptococcus mutans is composed of a hydrophilic domain, an LPNTGV motif, a hydrophobic domain, and a charged tail. These features are shared by surface proteins from many gram-positive coccal bacteria. To investigate the role of the C-terminal domains in antigen P1 surface localization, full-length and truncated P1 gene constructs, which were expressed on the shuttle vector pDL276, were transformed into the P1-negative mutant S. mutans SM3352, Streptococcus gordonii DL-1, and Enterococcus faecalis UV202. Transformants were tested for expression of P1 by enzyme-linked immunosorbent assaying and Western blotting. The results showed that full-length P1 was expressed by transformants of all three bacteria and was localized on the cell surface. A fusion protein composed of the Staphylococcus aureus fibronectin binding protein C terminus and the P1 protein N terminus was found to surface localize in S. mutans. Deletion of the entire C-terminal domains resulted in P1 being expressed in the culture supernatant. A P1 truncation, which carried only the hydrophilic domain at its C terminus, was found partially associated with the cell surface. This truncated P1 was readily removed from the isolated cell wall by hot sodium dodecyl sulfate-mercaptoethanol extraction. In contrast, the full-length P1 remained associated with the isolated cell wall after similar treatment, suggesting covalent linkages between the full-length P1 and the cell wall. The results described above showed that antigen P1 was anchored to the cell wall by its C-terminal domains probably via covalent linkages with the cell wall. The results also support a universal mechanism involving the C-terminal domains for protein surface localization among this group of gram-positive bacteria.

Isolation and partial characterization of uronic acid-containing glycoproteins from Mucor rouxii

Five different fractions containing uronic acids associated with protein were isolated from the cytoplasm of the filamentous form of Mucor rouxii. A single fraction was isolated from the cell wall by hot sodium dodecyl sulfate followed by ion exchange column chromatography. Two cytoplasmic entities (peaks I and II) were not adsorbed to DEAE Bio-Gel A. The molecular mass of peaks I to V ranged from 16.5 to 210 kDa. The protein-uronic acid ratios were different for each fraction. The cell wall fraction showed a molecular mass of 16.5 kDa, similar to that of peak II but with differences in chromatographic behavior and protein-uronic acid ratio. The possible role of these molecules as acceptors of sugar residues during polyuronide chain growth is discussed.

Targeting of a heterologous protein to the cell wall of Saccharomyces cerevisiae.

The sexual adhesion protein of Saccharomyces cerevisiae MAT alpha cells, alpha-agglutinin, could not be extracted from the cell wall with hot sodium dodecyl sulfate (SDS), but became soluble after digestion of the cell wall with laminarinase. This indicates that it is intimately associated with cell wall glucan. A fusion protein was constructed consisting of the signal sequence of yeast invertase, guar alpha-galactosidase, and the C-terminal half of the alpha-agglutinin. Most of the fusion protein was incorporated in the cell wall. A small amount could be extracted with SDS, but most of it could only be extracted with laminarinase. On the other hand, cells containing a construct consisting of the signal sequence of invertase and alpha-galactosidase released most of the alpha-galactosidase into the medium and all cell wall-associated alpha-galactosidase was released by SDS. Labelling with antibodies showed that the alpha-galactosidase part of the fusion protein was exposed on the surface of the cell wall. The results demonstrate that the C-terminal half of the alpha-agglutinin contains the information needed to incorporate a protein into the cell wall.

Unorthodox intrasubunit interactions in the cellulosome ofClostridium thermocellum

Clostridium thermocellum, an anaerobic thermophilic cellulolytic bacterium, produces an extremely cohesive, very high-molecular-mass multicellulase-containing complex termed thecellulosome. One of its components, the S1 subunit, is a nonenzymatic, 210 kDa glycopolypeptide. Upon preconditioning of the intact cellulosome with lowionic-strength or low-pH solutions, the S1 subunit separates in hot sodium dodecyl sulfate (SDS) solutions into a series of defined lowermolecular-mass subcomponents. Under the same conditions, the purified S1 subunit demonstrated the same behavior. Higher levels of glycosylation associated with the larger S1 subcomponents. The data support alterations in the conformational state of the S1 structure that lead to its disintegration induced by combined treatments with SDS and heating. Evidence is provided that this phenomenon may reflect a physiological response of the cellulosome, since similar alterations in S1 appear to accompany its binding to cellulose.

Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting analysis of smooth-lipopolysaccharide heterogeneity among Brucella biovars related to A and M specificities

Smooth (S)-lipopolysaccharide (LPS) preparations from reference and field strains of several biovars of Brucella abortus, B. melitensis, and B. suis were prepared by (i) the hot phenol-water method, (ii) hot sodium dodecyl sulfate extraction and proteinase K digestion, or (iii) dimethyl sulfoxide extraction. These S-LPS-enriched fractions were further analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and silver staining after periodate oxidation. Immunoblots were developed by using either monoclonal antibodies specific for Brucella A or M antigens or polyclonal polyspecific or monospecific sera from rabbits, cattle, and goats. The specificity of monoclonal antibodies reactive with Brucella unique (A or M) epitopes was demonstrated by enzyme-linked immunosorbent assay, LPS latex agglutination, or agglutination inhibition. The most-represented subunits of S-LPS ranged in Mr from 30,000 to 70,000 relative to marker proteins. According to A or M immunodominance, two sodium dodecyl sulfate-polyacrylamide gel electrophoresis banding patterns were clearly distinguished among biovars, whatever the fraction tested: a close succession of regularly spaced narrow bands for A greater than M strains and regularly spaced triplets of bands including either (i) a first thin band followed by two thick bands for B. abortus M greater than A strains or (ii) one thick band between two thin bands for B. melitensis or B. suis M greater than A strains. Moreover, A and M specificities were reaffirmed by sandwich enzyme immunoassay and latex agglutination inhibition with monoclonal antibodies and polyclonal sera.

Comparative immune responses to native cell envelope antigens and the hot sodium dodecyl sulfate insoluble fraction (PG) of Brucella abortus in cattle and mice

Brucella abortus vaccines composed of native cell envelopes or outer membrane proteins of smooth strain 2308 were compared with a vaccine (PG) composed of the insoluble residue of strain 2308 cell envelopes which had been extracted with hot sodium dodecyl sulfate. Vaccines were given by injection in an oil base adjuvant containing trehalose dimycolate and muramyl dipeptide or without adjuvant. Mice vaccinated with 30 μg native cell envelopes or PG and challenged 4 weeks later with virulent B. abortus strain 2308 displayed equivalent levels of protective immunity at 1 and 4 weeks post-infection. Heifers were vaccinated with 5 mg of antigens in adjuvant; PG was also administered without adjuvant. Humoral and cell mediated immune (CMI) responses were tested at monthly intervals. PG without adjuvant induced negligible immune responses. Native cell envelope antigens induced significantly higher titers of whole cell agglutinins over a 3-month period than did PG, although revaccination with PG in adjuvant enhanced the production of agglutinins and both vaccines induced antibodies to the O polysaccharide. Lymphocyte blastogenesis responses and delayed hypersensitivity reactions to porin and group 3 proteins were stimulated by both native and PG vaccines, and the magnitude of the responses did not differ significantly between the treatment groups. These vaccines were therefore comparable in their capacity to induce protective immunity in mice and CMI responses in cattle, whereas antibody responses induced by PG in cattle were generally lower. These findings provide a basis for evaluation of nonliving B. abortus vaccines in cattle.

The cell wall of Chlamydomonas reinhardtii gametes: Composition, structure and autolysin-mediated shedding and dissolution

The cell walls of Chlamydomonas gametes are multilayered structures supported on frameworks of polypeptides extending from the plasma membrane. The wall-polypeptide catalogue reported by Monk et al. (1983, Planta 158, 517-533) and extended by U.W. Goodenough et al. (1986, J. Cell Biol. 103, 405-417) was re-evaluated by comparative analysis of mechanically isolated cell walls purified from several strains. The extracellular locus of wall polypeptides was verified by in vivo iodogen-catalysed iodination and by autolysin-mediated elimination of the bulk of these polypeptides from the cell surface. Three (w15, w16, w17) and possibly four (w14) polypeptides were located to the most exterior aspect of the wall because of their susceptibility to Enzymobeadcatalysed iodination and their retention by a cell-wall-less mutant. The composition of shed walls stabilised with ethylenediaminetetraacetic acid during natural mating and kinetic analysis of the dissolution of walls purified from a bald-2 mutant demonstrated the rapid and specific destruction of polypeptide w3. Differential solubilisation of wall polypeptides occurred after loss of w3. Wall dissolution, characterised by the generation of fishbone structures from the W2 layer, gave as many as four additional polypeptides. Charged detergents and sodium perchlorate extracted a comparable range of polypeptides at room temperature from mechanically isolated walls, i.e. components of the W4-W6 layers, hot sodium dodecyl sulphate solubilised framework polypeptides, while reducing agent was required to solubilise the W2 layer. A model of wall structure is presented.

Structure of Bordetella pertussis peptidoglycan.

Bordetella pertussis Tohama phases I and III were grown to the late-exponential phase in liquid medium containing [3H]diaminopimelic acid and treated by a hot (96 degrees C) sodium dodecyl sulfate extraction procedure. Washed sodium dodecyl sulfate-insoluble residue from phases I and III consisted of complexes containing protein (ca. 40%) and peptidoglycan (60%). Subsequent treatment with proteinase K yielded purified peptidoglycan which contained N-acetylglucosamine, N-acetylmuramic acid, alanine, glutamic acid, and diaminopimelic acid in molar ratios of 1:1:2:1:1 and less than 2% protein. Radiochemical analyses indicated that 3H added in diaminopimelic acid was present in peptidoglycan-protein complexes and purified peptidoglycan as diaminopimelic acid exclusively and that pertussis peptidoglycan was not O acetylated, consistent with it being degraded completely by hen egg white lysozyme. Muramidase-derived disaccharide peptide monomers and peptide-cross-linked dimers and higher oligomers were isolated by molecular-sieve chromatography; from the distribution of these peptidoglycan fragments, the extent of peptide cross-linking of both phase I and III peptidoglycan was calculated to be ca. 48%. Unambiguous determination of the structure of muramidase-derived peptidoglycan fragments by fast atom bombardment-mass spectrometry and tandem mass spectrometry indicated that the pertussis peptidoglycan monomer fraction was surprisingly homogeneous, consisting of greater than 95% N-acetylglucosaminyl-N-acetylmuramyl-alanyl-glutamyl-diaminopimelyl++ +-alanine.

2-O-methyl-d-xylose containing sheath in the cyanobacterium Gloeothece sp. PCC 6501

The sheath of the unicellular cyanobacterium Gloeothece sp. PCC 6501 was isolated from cell homogenates by differential and sucrose gradient centrifugation, followed by lysozyme treatment and hot sodium dodecyl sulfate extraction. The sheath contains a major fraction of carbohydrate consisting of galactose, glucose, mannose, rhamnose, 2-O-methyl-d-xylose, xylose, glucuronic and galacturonic acids, but only traces of fatty acids and phosphate. A protein content of about 2% (of fraction dry weight) could not be removed by the detergent treatment.