Position‐Dependent Electrostatic Protection against Protein Aggregation

Proteins with a high propensity to aggregate can be largely prevented from doing so with surprisingly small changes to their primary structure. By using a combination of rational design and quantitative measurements of aggregation rates, we show that adding a single charge in specific “gatekeeper” regions is sufficient to change the timescale for amyloid fibril growth from minutes to weeks, thereby dramatically reducing the efficiency of this process.

     

Determination of NAPL-water interfacial areas in well-characterized porous media

The nonaqueous-phase liquid (NAPL)-water interfacial area is an important parameter which influences the rate of NAPL dissolution in porous media. The aim of this study was to generate a set of baseline data for specific interfacial area for a two-phase-entrapped NAPL-water system in well-characterized porous media and subsequently use these data to evaluate two current theoretical models. The first model tested distributes entrapped NAPL over the pore classes based on Land's algorithm and assumes the resulting blobs to be spherical. The other model is thermodynamically based, assuming that reversible work done on the system results in an increase in interfacial area, such that the area between drainage and imbibition retention curves can be related to the interfacial area. Interfacial tracer tests (IFTT) were used to measure specific entrapped NAPL (hexadecane)-water interfacial areas in columns packed with four grades (12/20, 20/30, 30/40, 40/50) of silica sand. By use of the anionic surfactant dihexylsulfosuccinate (Aerosol MA80), IFTT gave specific interfacial areas between 58 cm(-1) for the finest sand and 16 cm(-1) for the coarsest, compared to values of between 33 and 7 cm(-1) for the first model and between 19 and 5 cm(-1) for the thermodynamic model. Results from the literature suggest that nonspherical blobs shapes occur relatively frequently; hence it is reasonable to suggest that the assumption of spherical NAPL blobs may explain the underprediction by the first model. The thermodynamic model underestimates the interfacial area because it assumes that entrapment occurs only within the largest pores. A modified version of the latter model, allowing entrapment across all pore classes, yielded values between 58 and 13 cm(-1). Of the models tested the modified thermodynamic model best predicts the interfacial area.     

Principles and design of a novel magnetic force mechanical conditioning bioreactor for tissue engineering, stem cell conditioning, and dynamic in vitro screening

Mechanical conditioning of cells and tissue constructs in bioreactors is an important factor in determining the properties of tissue being produced. Mechanical conditioning within a bioreactor environment, however, has proven difficult. This paper presents the theoretical basis, design, and initial results of a mechanical conditioning system for cell and tissue culture which is based on biocompatible magnetic micro- and nanoparticles acting as a remote stress mechanism without invasion of the sterile bioreactor environment.     

Real-time PCR detection of bacteria belonging to the Firmicutes Phylum

Members of the bacterial Phylum Firmicutes occupy a wide range of habitats and can be either beneficial or detrimental in diverse settings, including food- and beverage-related industries. Firmicutes are responsible for the vast majority of beer-spoilage incidents and, as such, they have a substantial financial impact in the brewing industry. Rapid detection and identification of a bacterium as a Firmicutes is difficult due to widespread genetic transfer and genome reduction resulting in phenotypic diversity in these bacteria. Here we describe a real-time multiplex PCR to detect and differentiate Firmicutes associated with beer-spoilage from non-Firmicutes bacteria that may be present as benign environmental contaminants. A region of the 16S rRNA gene was identified and predicted to be highly conserved amongst, and essentially specific for, Firmicutes. A real-time PCR assay using a hydrolysis probe targeting this region of the 16S rRNA gene was experimentally shown to detect ten genera of Firmicutes known to be beer spoilers, but does not cross-react with eleven of twelve non-Firmicutes genera which can periodically appear in beer. Only one non-Firmicutes species, Zymomonas mobilis, weakly reacted with the Firmicutes probe. This rPCR assay has a standard curve that is linear over six orders of magnitude of DNA, with a quantitation limit of DNA from <10 bacteria. When used to detect bacteria present in beer, the assay was able to detect 50-100 colony forming units (CFU) of Firmicutes directly from 2.5 cm membranes used to filter 100 ml of contaminated beer. Through incorporation of a 4.7 cm filter and an overnight pre-enrichment incubation, the sensitivity was increased to 2.5-10 CFU per package of beer (341 ml). When multiplexed with a second hydrolysis probe targeting a universal region of the 16S rRNA gene, the assay reliably differentiates between Firmicutes and non-Firmicutes bacteria found in breweries.     

Identification and treatability of organics in oil shale retort water

Analysis by GC-MS of the process water derived from Fischer Assay retoring of oil shale from Rundle, Australia has provided positive identification of the major organic constituents present. This is the first detailed analysis of retort water from Australian oil shales and showed that the compounds ranged from being highly biodegradable to highly inhibitory and resistant to biological oxidation. The major classes of compounds found in a composited sample included normal carboxylic acids, alkyl pyridines, quinolines and cyclic saturated and unsaturated ketones. Separation of the retort water into its acid, base and neutral fractions was brought about by solvent extraction using methylene chloride. A series of treatability studies on the retort water confirmed the hypothesis that only a portion of the organic carbon was amenable to biological treatment. In addition, high ammonia levels further inhibited biological action. Adsorption of the retort water with activated carbon proved most successful in removing the non-biodegradable fraction of the organic species. Chemical oxidation by ozone does not appear attractive because it lacks the specificity of adsorption.     

Characteristics and properties of electrodeposited chromium from aqueous solutions

The characteristics and properties of chromium deposited from sulphate-catalysed chromic acid solutions of varying composition over both a current density and temperature range have been examined. An attempt to interpret the mechanical test results on the basis of inherent crack incidence and crack length within the deposit has also been presented.Comparisons between laboratory and commercial platings have established the viability of extrapolating the laboratory results, enabling test gun tube performance to be related to the various coating structures produced in the 120 mm barrel by the varying current density conditions experienced. These current density variations have been predicted using the boundary element numerical method which demonstrates the inherent problems associated with gun tube geometry.This paper was presented at a workshop on the electrodeposition of refractory metals, held at Imperial College, London, in July 1985.     

Conformation-activity correlations for chemotactic tripeptide analogs incorporating dialkyl residues with linear and cyclic alkyl sidechains at position 2

Five stereochemically constrained analogs of the chemotactic tripeptide incorporating 1-aminocycloalkane-1-carboxylic acid (Ac(n)c) and alpha,alpha-dialkylglycines (Deg, diethylglycine; Dpg, n,n-dipropylglycine and Dbg, n,n-dibutylglycine) at position 2 have been synthesized. NMR studies of peptides For-Met-Xxx-Phe-OMe (Xxx=Ac(7)c, I; Ac(8)c, II; Deg, III; Dpg, IV and Dbg, V; For, formyl) establish that peptides with cycloalkyl residues, I and II, adopt folded beta-turn conformations in CDCl3 and (CD3)2SO. In contrast, analogs with linear alkyl sidechains, III-V, favour fully extended (C5) conformations in solution. Peptides I-V exhibit high activity in inducing beta-glucosaminidase release from rabbit neutrophils, with ED50 values ranging from 1.4-8.0 x 10(-11) M. In human neutrophils the Dxg peptides III-V have ED50 values ranging from 2.3 x 10(-8) to 5.9 x 10(-10) M, with the activity order being V > IV > III. While peptides I-IV are less active than the parent, For-Met-Leu-Phe-OH, in stimulating histamine release from human basophils, the Dbg peptide V is appreciably more potent, suggesting its potential utility as a probe for formyl peptide receptors.     

Needlescopic adrenalectomy--the initial series: comparison with conventional laparoscopic adrenalectomy

Functional group gradients were prepared on low-density polyethylene (PE) sheets. The surface density of grafted functional groups was gradually changed along the sample length by way of corona discharge treatment with gradually increasing power following graft copolymerization of acrylic acid (AA), sodium p-styrene sulfonate (NaSS), or N,N-dimethyl aminopropyl acrylamide (DMAPAA). AA and NaSS are negatively chargeable and DMAPAA is positively chargeable in phosphate-buffered saline or plasma solution at pH 7.3-7.4. The prepared functional group gradient surfaces were characterized by measurement of the water contact angle, by electron spectroscopy for chemical analysis, and by Fourier transform infrared spectroscopy in the attenuated total reflectance mode. All these measurements indicated that the functional groups were grafted onto the PE surfaces with gradually increasing density. The platelets adhered to the functional group gradient surfaces along the sample length were counted and observed by scanning electron microscopy. It was observed that the platelet adhesion to the gradient surfaces decreased gradually with the increasing surface density of functional groups. This may be related to the hydrophilicity of the surfaces. The DMAPAA-grafted surface showed a large amount of platelet adhesion, probably due to its positive charge character, while the AA-grafted surface, which is charged negatively, showed poor platelet adhesion. However, the NaSS-grafted surface, which is also charged negatively, showed a relatively large amount of platelet adhesion. This may be associated with the existence of an aromatic ring close to the ionizable group in NaSS. It seems that surface functional groups and their charge character, as well as wettability, play important roles for platelet adhesion.