Effects of eucaryotic initiation factor 3 on eucaryotic ribosomal subunit equilibrium and kinetics

In order to understand the possible role of eucaryotic initiator factor 3 (eIF-3) in maintaining a pool of eucaryotic subunits, we have measured the effects of eIF-3 on the equilibria and kinetics of ribosomal subunit association and dissociation. The ribosomal subunit interactions have been studied by laser light scattering, which does not perturb the system. We find that eIF-3 reduces the apparent association rate of reticulocyte, wheat germ, and Artemia ribosomes. The kinetics of the reassociation for a shift in [Mg2+] from 0.5 to 6 mM are best explained by a model where eIF-3 dissociates from the 40S subunits prior to association of the 40S and 60S subunits. Static titrations indicate there is some binding of eIF-3 to 80S ribosomes at lower [Mg2+].     

Influence of ion gradients on the transbilayer distribution of dibucaine in large unilamellar vesicles

The uptake of dibucaine into large unilamellar vesicles in response to proton gradients (delta pH; inside acidic) or membrane potentials (delta psi; inside negative) has been investigated. Dibucaine uptake in response to delta pH proceeds rapidly in a manner consistent with permeation of the neutral (deprotonated) form of the drug, reaching a Henderson-Hasselbach equilibrium where [dibucaine]in/[dibucaine]out = [H+]in/[H+]out and where the absolute amount of drug accumulated is sensitive to the buffering capacity of the interior environment. Under appropriate conditions, high absolute interior concentrations of the drug can be achieved (approximately 120 mM) in combination with high trapping efficiencies (in excess of 90%). Dibucaine uptake in response to delta psi proceeds more than an order of magnitude more slowly and cannot be directly attributed to uptake in response to the delta pH induced by delta psi. This induced delta pH is too small (less than or equal to 1.5 pH units) to account for the transmembrane dibucaine concentration gradients achieved and does not come to electrochemical equilibrium with delta psi. Results supporting the possibility that the charged (protonated) form of dibucaine can be accumulated in response to delta psi were obtained by employing a permanently positively charged dibucaine analogue (N-methyldibucaine). Further, the results suggest that delta psi-dependent uptake may depend on formation of a precipitate of the drug in the vesicle interior. The uptake of dibucaine into vesicles in response to ion gradients is of direct utility in drug delivery and controlled release applications and is related to processes of drug sequestration by cells and organelles in vivo.     

Sequential modification of membrane currents with classical conditioning

Pavlovian conditioning of the nudibranch mollusc Hermissenda crassicornis was previously shown to produce long-lasting reduction of two K+ currents measured across the Type B photoreceptor soma membrane (Alkon et al., 1982a; Alkon et al., 1985). Pavlovian conditioning of the rabbit was also shown to be followed by persistent K+ current reduction (Disterhoft et al., 1986). Here we report the first evidence that Ca2+ currents can also be modified by conditioning. The amplitude of the currents rather than their voltage-dependence remains reduced at least 1-2 d after conditioning (but not control procedures). Conditioning-induced changes of both K+ and Ca2+ currents increased as a function of training, the Ca2+ currents only changing substantially with greater than or equal to 250 trials. The later changes of the Ca2+ current may function to limit the magnitude of excitability increases due to associative learning.     

Reactions of d-glyceraldehyde 3-phosphate dehydrogenase with chromophoric thiol reagents

The kinetics of the reaction of d-glyceraldehyde 3-phosphate dehydrogenase with 5,5'-dithiobis-(2-nitrobenzoic acid) show that NAD(+) dissociates from the enzyme before the reaction. In contrast 2-chloromercuri-4-nitrophenol reacts with the holoenzyme without prior dissociation of NAD(+). These studies and observations on the dissociation constant of NAD(+) to the lobster enzyme show that NAD(+) must dissociate from sites modified by substrates during the reductive dephosphorylation of 1,3-diphosphoglycerate. All four sites per tetramer of the apoenzyme are acylated by 1,3-diphosphoglycerate. Hydrolysis of the acyl-enzyme occurs at a significant rate even in the absence of NAD(+), which may explain previous estimates that only two sites per tetramer can readily be acylated.     

The fate of the fusarium mycotoxin zearalenone in maize cell suspension cultures

The fusarium mycotoxin zearalenone was transformed in cell suspension cultures of Zea mays giving α- and β-zearalenol and the β-D-glu cos ides of zearalenone and α- and β-zearalenol. The structure of zearalenone-4-β-D-glucopyranoside was determined by liquid — chromatography-mass spectrometry and specific hydrolysis with β-glucosidase. α- and β-zearalenol and their glucosides were identified by co chromatography using tic and HPLC and glucosidase — treatment Up to 50% of the mycotoxin added was bound to a non extractable or “bound” residue fraction. After treating this residue by a sequential cell wall fractionation procedure, zearalenone was found to be bound mainly to starch, hemicellulose, and lignin fractions.     

Gene transfer is a major factor in bacterial evolution

Lateral gene transfer in four strains of Salmonella enterica has been assessed using genomic subtraction. Strain LT2 (subspecies I serovar Typhimurium) chromosomal DNA was used as target and subtracted by three subspecies I strains of serovars Typhimurium (S21), Muenchen (S71), Typhi (M229), and a subspecies V strain (M321). Data from probing random cosmids of LT2 DNA with preparations of the residual LT2 DNA after subtraction were used to estimate the amounts of LT2 DNA not able to hybridize to strains S21, S71, M229, and M321 to be in the range of 84-106, 191-355, 305-629, and 778-1,286 kb, respectively. Several lines of evidence indicate that most of this DNA is from genes not present in strain M321 and not from genes that have diverged in sequence. The amounts correlate with the divergence of the four strains as revealed by multilocus enzyme electrophoresis and sequence variation of housekeeping genes. Sequence of 39 of the fragments from the M321 subtracted residual LT2 DNA revealed only six inserts of known gene function with evidence of both gain and loss of genes during the development of S. enterica clones. Sixteen of the 39 segments have 45% or lower G+C content, below the species average, but over half are within the normal range for the species. We conclude that even within a species, clones may differ by up to 20% of chromosomal DNA, indicating a major role for lateral transfer, and that on the basis of G+C content, a significant proportion of the DNA is from distantly related species.      

Interaction of chemosensory, visual, and statocyst pathways in Hermissenda crassicornis

Neurons in the cerebropleural ganglia (CPG), photoreceptors in the eye, optic ganglion cells, and statocyst hair cells of the nudibranch mollusk Hermissenda crassicornis responded in specific ways, as recorded intracellularly, to stimulation of the chemosensory pathway originating at the tentacular chemoreceptors as well as to stimulation of the visual pathway originating at the photoreceptors. Synaptic inhibition of photoreceptors occurs via the chemosensory pathway. The possible significance of such intersensory interaction is discussed with reference to preliminary investigation of the animal's gustatory behavior and possible neural mechanisms of behavioral choice.     

A finite element study of the effect of diametral interface gaps on the contact areas and pressures in uncemented cylindrical femoral total hip components

Uncemented femoral total hip components rely entirely on contact with the prepared femur for their initial fixation. The contact areas and stresses between a straight tubular bone and a metal cylindrical prosthesis 12.5 cm long and 13 mm in diameter were calculated in a finite element model which includes uniform diametral gaps varying from 20 to 500 microns, using transverse loads from 100 to 2000 N. Frictionless three-dimensional contact elements were used between the bone and the prosthesis. Contact stresses were high and irregular in all cases, and the contact areas were small. Two regions of contact were apparent for lower loads and larger gaps. A third region of contact occurred near the distal tip of the implant at higher loads. This region of contact markedly increased the contact stresses at the distal tip of the prosthesis. A 20 microns overlap between bone and implant was modelled to assess a slight interference fit. The contact stress distribution in this case was markedly different from the stress distribution with a 20 microns diametral gap. The data collectively indicates that gaps of less than 20 microns between bone and implant can substantially change contact stress distributions.     

An analytical and numerical study of the stability of bone remodelling theories: dependence on microstructural stimulus.

The origin of unstable bone remodelling simulations using strain-energy-based remodelling rules was studied mathematically in order to assess whether the unstable behavior was due to the mathematical rules proposed to characterize the processes, or to the numerical approximations used to exercise the mathematical predictions. A condition which is necessary for the stability of a strain-energy-based remodelling theory was derived analytically using the calculus of variation. The analytical result was derived using a simple elastic model which consists of a long beam loaded by an axial force and a bending moment. This loading situation mimics the coupling between local density and global density distributions seen in vivo. A condition necessary for a stable remodelling scheme is arrived at, but the conditions necessary to guarantee a stable remodelling scheme are not. In this remodelling scheme, the elastic modulus is proportional to volumetric density raised to an exponent n, and the microstructural stimulus is taken as the strain energy density divided by volumetric density raised to an exponent m. In order for a remodelling scheme to be stable in this loading situation, m must be greater than n. Finite-difference time-stepping is used to verify the predictions of the analytical study. These numerical studies appear to confirm the analytical studies. Physiologic interpretation of the behavior found with n greater than m indicates that this type of unstable behavior is unlikely to be observed in vivo. Since numerical approximations are not made in deriving this stability condition, we conclude that the mathematical rules proposed to characterize bone remodelling based on strain energy density should meet this condition to be relevant to physiologic bone remodelling.