Sequence requirements for cytochromes P450IIA1 and P450IIA2 catalytic activity: evidence for both specific and non-specific substrate binding interactions through use of chimeric cDNAs and cDNA expression

Cytochrome P450s IIA1 and IIA2, encoded by the CYP2A1 and CYP2A2genes, display 88% amino acid sequence similarities. The dissimilaritiesof sequence between these two enzymes are primarily localizedwithin four discrete regions of the polypeptides that are separatedby regions of absolute sequence identity. IIA1 specificallyhydroxylates the prototype substrate testosterone at the 7 and6 position with a predominance of 7 metabolite. IIA2, on theother hand, hydroxylates this steroid at eight positions onthe molecule, with one of the most abundant metabolites being15hydroxytestosterone. To determine those amino acids responsiblefor the difference in testosterone hydroxylation specificities,chimeras were constructed between IIA1 and IIA2 cDNAs and expressedin cell culture using vaccinia-virus-mediated cDNA expression.Chimeras, in which the first 355 amino acids correspond to asingle enzyme, maintain the specificity associated with thatenzyme. Of six chimeras which have substitutions between aminoacids 161 and 276, two are inactive and the remaining four givesimilar metabolite profiles, in which both 7 and 15 hydroxylationspecificities have been lost. Two of these four chimeras arediametric apposites, suggesting that modification of eitherthe N-terminal or central regions of the enzymes results inconformational changes that prevent the specific binding interactionsresponsible for the narrow regioselectivity associated withIIA1 and 15-hydroxytestosterone formation associated with IIA2.     

Estimation of Mechanical Parameters in Multicompartment Models Applied to Normal and Obstructed Lungs During Tidal Breathing

A technique is presented which allows quantitative assessment of the use of parallel compartment models for characterizing pulmonary mechanical function during tidal breathing. A model consisting of a conducting airway leading to two parallel parenchymal regions is used. Numerical simulation and sensitivity analysis indicated that a) the compliance of the conducting airway was not significant under the experimental conditions of interest and that b) estimates of the distribution of central and peripheral resistances would not be precise. The techniques were demonstrated using measurements of transpulmonary pressure, flow, and volume changes during tidal breathing obtained from a human subject with normal lungs and a human subject with obstructed lungs. Optimal estimates of the parameters were obtained by minimizing the difference between the model output and experimental data combined from two breathing frequencies. In the estimation procedure, the sum of the peripheral compliances was constrained to equal the independently measured static lung compliance. This constraint was critical for correct evaluation of nonuniform mechanical lung function. From the parameter estimates, the ratio of parenchymal time constants was about five in the subject with normal lungs and 60 in the subject with obstructed lungs. These results suggest that a full study with several normal and obstructed lung subjects is warranted.     

Statistical Measures of Parameter Estimates from Models Fit to Respiratory Impedance Data: Emphasis on Joint Vanabilities

To describe respiratory mechanical impedance data, many investigators have proposed electromechanical models and then fit them to data using formal parameter estimation techniques. This approach has resulted in confusion as to how to interpret the resulting estimated values, and hence as to which model is most appropriate. A key cause of this confusion is that most studies rely on the quality of fit between the model and the data as the only measure of model validity rather than performing adequate statistical analysis of the parameter estimates themselves. This paper describes several statistical measures that should be applied to parameter estimates obtained from forced oscillation data. Specifically, we describe standard errors of the parameter estimates, confidence intervals for each parameter estimate, and the joint confidence region for the parameters. Much emphasis is placed on the joint confidence region which, unlike the interval, allows for simultaneous variations in parameters. The measures are applied to an often used six-element model for respiratory impedance data of dogs from 4 to 64 Hz. This application indicated that even when fitting data over this frequency range, parameter estimates are not well defined and the parameter estimated with least accuracy is airway resistance.     

Solid-State Electrodes for Multichannel Multiplexed Intracortical Neuronal Recording

Thin-film arrays of extracellular recording electrodes have been developed for use in studies of information processing in neural structures and eventual use in closed-loop control of neural prostheses. These probes consist of a silicon substrate which supports an array of thin-film conductors. The conductors are insulated above and below with deposited dielectrics. The electrode sites are defined by openings in the upper dielectric layer and are inlaid with gold to form low-impedance recording surfaces. The probes are typically 15 pim in thickness with shank widths as narrow as 20 ?m. The probe fabrication process is compatible with the inclusion of signal processing circuitry directly on the probe substrate. A 12 channel on-chip signal processor design with per-channel gain of 100, bandwidth of 100 Hz-6 kHz, multiplexed output, and recording-site impedance check capability is described. The probes have adequate strength to penetrate the gerbil pia-arachnoid layer and have recorded single neuron activity of over 500 ?V peak-to-peak from tip, side, and mid-carrier sites. Signal-to-noise ratios as high as 10:1 have been achieved. An equivalent circuit model for the conducting leads, the recording site, and the electrode-electrolyte interface is described. Development of biocompatible insulation and encapsulation materials for long-term implantation of active probes is underway.     

Direct and Indirect Dissolution of Sapphire in Calcia–Magnesia–Alumina–Silica Melts: Dissolution Kinetics

The dissolution rate of sapphire into CaO-MgO-Al₂O₃-SiO₂ (CMAS) melts was investigated at 1450° and 1550°C. The effects of immersion time, specimen rotation rate, and magnesia concentration in the bulk melt on the sapphire dissolution rate and on the growth rate of magnesium aluminate spinel at the sapphire/melt interface were determined. The processes of direct and indirect sapphire dissolution into CMAS melts are discussed, with the support of kinetic and microprobe data.¹ A model of indirect sapphire dissolution^1,2 is used along with the equations of Tedmon³ to describe the kinetics of indirect dissolution and spinel layer growth during forced-convective and static sapphire dissolution.     

Compressive Creep of SiC-Whisker-Reinforced Al2O3

Compressive creep of SiC-whisker-reinforced Al₂O₃ composites (0, 5, 15, and 25 wt% SiC) was measured in the temperature range of 1300° to 1500°C in air and argon. The creep resistance increased with increasing whisker concentration. The results indicated that the whiskers degraded in air, increasing strain rates compared to those in argon. Stress exponents between 1.0 and 2.0 and an activation energy of 620 ± 100 kJ/mol were measured. Transmission electron microscopy observations indicated that cavitation was minimal and that the deformed composites had the same dislocation structure as did the as-received samples.     

Autoxidation of methyl linolenate and methyl linoleate: The effect of α-tocopherol

α-Tocopherol substantially affects the distribution of peroxidic compounds formed during the autoxidation of methyl linolenate and methyl linoleate. In the autoxidation of both these unsaturated fatty acid esters the proportion of monohydroperoxides with conjugated double bonds in the trans, trans configuration is reduced until at high concentrations of α-tocopherol (5%) only cis, trans isomers are formed. In the autoxidation of methyl linolenate the proportion of hydroperoxy-epidioxides is reduced and only monohydroperoxides are formed when 5% α-tocopherol is present. The results are discussed in terms of recent findings concerning the mechanism of the autoxidation of unsaturated lipids.     

Analysis of the behavior of transverse walls of socket base connections

This paper presents a theoretical and experimental analysis of socket base connections of precast concrete structures with regard to the behavior of transverse walls. The experimental program included seven specimens, for which the type of interface in contact with cast-in-place concrete, the load eccentricities and the embedded lengths were all varied. A design model was proposed to calculate the reinforcements of the transverse walls. Based on the obtained results, some conclusions can be drawn: (a) The top of the transverse wall on the compression side of the smooth connections and the top of the two transverse walls of the rough connections are submitted to a bending-tension and this tension prevails on the bending; (b) The design model proposed for the calculation of the reinforcement of the transverse wall on the compression side provides the best prediction of the experimental results for all specimens when compared to the current design models; (c) For rough interfaces, the top of the transverse wall on the tension side is more requested than the top of transverse wall on the compression side; (d) The results of the proposed design model for the reinforcement of the transverse wall on the tension side of rough connections were in close agreement with the experimental results.     

Analytical models for the series resistance of selective emitters in silicon solar cells including the effect of busbars

A theoretical analysis of the power loss and series resistance of the front side emitter in silicon solar cells is presented. Existing 1D models (infinitely long finger) and 2D models (including the effect of busbars) of emitter series resistance contribution are extended to the case of selective emitters. The general case of different current densities for both emitters in the selective emitter scheme is considered in these extensions. The resulting models depend on the individual sheet resistances and current densities in both emitters and the device's overall grid geometry. The models are corroborated by finite element simulation of the potential in the emitter. An excellent agreement is found between the analytical models, and the simulations for a wide range of sheet resistances typically encountered in silicon solar cells. Grid simulations using the 2D model are applied to solar cells with selective emitters, where the width of the low‐resistive emitter was varied. The simulations demonstrate that the 2D model can explain the absolute change in fill factor observed in these cells. Copyright © 2013 John Wiley & Sons, Ltd.