A new approach for the purification and characterisation of PA49.5, the main prebiotic of Lactococcus lactis subsp. cremoris

The main autolysin PA49.5, an enzyme that hydrolyzes or destroys the components of a biological endogenous cell or a tissue, was purified 3045 times from the homogenate of a whole cell extract of Lactococcus lactis subsp. cremoris ATCC 9596 (Mc5), with a recovery yield of 52%. The purification of the protein was carried out through a micro-purification technique using SDS-BigCHAP polyacrylamide gel electrophoresis and concentrated with a Microcon-10 filtration system. SDS-polyacrylamide gel electrophoresis of the purified enzyme confirmed the presence of only one band having a molecular weight of 49.5 kDa. In view of its insolubility, PA49.5 contained in the cell extract precipitate was solubilized in the presence of 0.1% (w/v) of BigCHAP, a non-ionic detergent. Higher concentrations of this detergent completely inhibited the activity of solubilized PA49.5 or prevented its solubilization. The optimal pH and temperature for PA49.5 enzymatic activity are 7.5 and 45 degrees C respectively. In addition 0.1% or less of PA49.5 significantly increased Mc5 lysis. We observed 55% more lysis with 0.25 mug of purified PA49.5 compared to the control. Gas chromatography analysis of the components of the crude cell extract, of the precipitate and of the supernatant indicates the presence of at least 6 fatty acids. The long-chained fatty acids (e.g. C18:0 and C18:3) detected represent 81.65% of the precipitate from which PA49.5 was purified. Of these two acids, the C18:0 (stearic acid) alone represents 47.40% of the precipitate. Mc5 releases proteins at the beginning (major peak) and at the end (moderate peak) of the exponential stage of growth. Analysis by denaturing polyacrylamide gel electrophoresis with Mc5 cell walls incorporated as the autolysin's substrate identified a band corresponding to PA49.5 in the second peak of protein secretion.     

Conformational change due to esterification of hydroxy groups in erythromycin A and its major metabolite: analysis of these derivatives with different biological properties using NMR and molecular dynamics (MD) data

A conformational study is performed on the acylated erythromycin and erythralosamine derivatives from comparison between experimental results (NMR) and theoretical calculations by Molecular Dynamics (MD) in attempts to correlate their conformations with their abilities to generate cytochrome P450-nitroso metabolite complexes in vitro. As the 3'-dimethyl-amino function of the desosamine is metabolized and responsible for the interaction with cytochrome P450, its position, mobility and steric hindrance in the proximity of this functional group are related to its biological properties. The major conformations of the lactone ring were termed A (A1, A2, A3) and B (B1, B2), and this macrocycle flexibility induced five different orientations a, b, c, d and e for the desosamine sugar. Conformations A and B differ in many ways but the major change is the inward folding of the C(3) fragment in B. Conformer a exhibits an orientation of the desosamine nearly perpendicular to the macrocycle whereas the two units are in the same plane in conformations c and e. For conformation b, the cladinose unit lifts up above the macrocycle. Conformation d exhibits a turned-back cladinose. In the erythromycin derivatives esterification at the beta position to the N(CH3)2 group of the desosamine reduces the degree of freedom of the macrocyclic lactone ring which corresponds to conformation A only. The desosamine sugar was found to be perpendicular to the macrocycle (a conformer) and both sugar groups are parallel to reduce the steric energy. In the erythralosamine derivatives, the macrocycle is always present as conformation B with the two conformations b and c of the sugar rings. The steric parameters favour the b conformers in which the amino group is tilted up, while in 3,2'-dibenzoylated stacking aromatic attraction stabilizes the planar c conformer. Both isomers are thus shown to adopt well-defined conformations and to be well-adapted for a comparative structure-activity correlation studies. There is a significant relationship between the conformation b and the formation of cytochrome P450-nitroso metabolite complexes.     

Set point trajectory and internal force control in redundant dual-arm manipulation

A nonlinear decoupling and linearizing feedback control is considered for dynamic coordination of two planar robot arms manipulating an object. A general inverse dynamics-based method is presented that assures an exact feedback linearization for simultaneous control of the object trajectory on the plane and internal efforts transmitted from the robot end-effectors to the object. The method takes the manipulator dynamics and object dynamics into consideration. A method for parameterizing the grip matrix null space is proposed, which has formed a basis for developing a new method for calculating the internal efforts. The procedure is invariant with respect to the change of the torque origin and units of length, and provides the force distribution without internal squeezing effects. A comparison between the approaches known so far and the new method is presented. No previously published method assures noninvariance and nonsqueezing properties for all possible contact configurations. Control algorithms are developed for a system of robotic arms that has more degrees of freedom than necessary for given tasks, exhibiting both actuation and kinematic redundancy. The implementation of this method is demonstrated for the case of a system of two planar three-link arms with the end-effectors manipulating an object, with different constrained task configurations. Practical aspects of discrete-time inverse dynamics control, such as influence of the computational time delay and robustness to model imperfections, are discussed. It is demonstrated that it is possible to achieve high-precision tracking of object position and internal force profiles, even if a system imperfect model is used for controller design. © 1993 John Wiley & Sons, Inc.     

Processing speed in the 1st year of life: A longitudinal study of preterm and full-term infants.

Processing speed was assessed at 5, 7, and 12 months in full-term and preterm infants (birthweight     

Room-temperature diffusion in Cu/Ag thin-film couples caused by anodic dissolution

Thin, 100-nm films of first silver and then copper were deposited consecutively onto inert substrates by magnetron sputter deposition. Constant anodic current densities were applied at room temperature to dissolve the outer copper film to varying depths. The 50Cu/50Ag interface, derived from the auger electron spectroscopic concentration-depth profile, initially moved into the copper toward the outer dissolving surface, indicating enhanced diffusion of copper into silver. After longer times at all anodic current densities, the interface reversed and moved back toward the underlying silver-rich layer, indicating that eventually diffusion of silver into copper predominated. The reversal time was inversely proportional to the anodic current density. These effects are explained by anodic formation of subsurface vacancies which migrate as divacancies to the copper/silver interface where they affect interface movements by the well-known Kirkendall mechanism. Calculated diffusivities up to 10⁻¹² cm²/s at maximum anodic current densities of 900 μA/cm² are dramatically above any that are normally observed at room temperature.     

Modeling of two-dimensional random fields

This paper presents a method of conditional stochastic modeling of two-dimensional fields which can be used to predict values at certain field points at a given time, based on field values at other locations at the same time and on data about second order field moments at given points. For computer simulations, the Gaussian truncated distributions are used. The aim of this work is also to present a derivation of a formula for the probability density of an n-dimensional random variable with the Gaussian conditional truncated distribution. As a numerical example, a soil contamination field described by correlation functions corresponding to the white noise field, the Shinozuka field and the Markov field is analyzed. The acceptance-rejection method is applied to generate covariance matrices and vectors of field values. Then, conditional expected field values for adequate correlation functions are calculated.     

Elastin-derived peptides and anti-elastic antibodies in children with recurrent or obstructive bronchitis

The aim of the study was to examine the effects of bronchitis on elastin-derived peptides (EP) and antielastin antibodies (AEAb) in serum. Using an ELISA technique, these parameters were assessed in children aged 6 months to 15 years: 36 had recurrent bronchitis (RB), 34 had obstructive bronchitis (OB) and 37 were healthy children (C). A comparison of the levels of the investigated compounds in the acute phase (I) and stage without signs of diseases (II) was studied. The EP data show non-significant changes in the RB and OB groups in comparison with group C. Children with OB had significantly lower levels of AEAb in IgA, IgG and IgM class antibodies, while those with RB in the IgG class in comparison with group C in both examinations. In examination 1, AEAb in IgA and IgM classes were significantly lower in children with OB than with RB.     

Numerical methods for Orr–Sommerfeld problems

Five previously employed numerical methods for the solution of Orr–Sommerfeld problems have been compared to each other and to a new method, the differential method of near-orthonormalized integration. Brief summaries of each method are included. The comparison, based on seven factors, reflects the results of an implementation of a computer program for each method for the classic Orr–Sommerfeld problem of plane Poiseuille flow. This comparison shows that the new method and the algebraic finite difference method are currently the best available numerical solution methods for the problems in this class, with the new method being less problem dependent.