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Biomedical subjects

J Frank

Publications and source records attributed to J Frank.

At least 271 records · Page 15Linked to original sources
PubMed1990

Three-dimensional reconstruction of native Androctonus australis hemocyanin.

A sample of native 4 x 6-meric hemocyanin of Androctonus australis was negatively stained with the double-layer technique, and was observed by transmission electron microscopy under low-dose conditions with a 50 degree and 0 degree tilt. The three-dimensional reconstruction method from "Single-exposure, random conical tilt series" was then applied. Independent three-dimensional reconstructions were obtained from the top, side and 45 degree views. Despite a pronounced flattening effect, presumably due to the specimen preparation technique, the positions of the 24 subunits composing the oligomer were unequivocally determined. This experiment definitely solves the problem of the architectural organization of the subunits in the cheliceratan 4 x 6-meric hemocyanins. Moreover, distinction between the flip and flop faces and an attenuated rocking effect were observed.

Animals↗
PubMed1990

Isolation and characterization of cytochrome c550 from the methylamine-oxidizing electron-transport chain of Thiobacillus versutus.

The isolation and purification of cytochrome c550 from the methylamine-oxidizing electron-transport chain in Thiobacillus versutus is reported. The cytochrome is a single-heme-containing type I cytochrome c with a relative molecular mass of 16 +/- 1 kDa, an isoelectric point of 4.6 +/- 0.1, a midpoint potential of 272 +/- 3 mV at pH less than 4 and 255 +/- 5 mV at pH = 7.0, and an axial coordination of the Fe by a methionine and a histidine. The midpoint potential decreases with increasing pH due to the deprotonation of a group tentatively identified as a propionate (pKa = 6.5 +/- 0.1 and 6.7 +/- 0.1 in the oxidized and reduced protein, respectively) and a change in the Fe coordination at pH greater than 10. The electron-self-exchange rate appears to depend strongly on the ionic strength of the solution and is relatively insensitive to changes in pH. At 313 K and pH 5.2 the electron-exchange rate amounts to 0.7 x 10(2) M-1 s-1 and 5.3 x 10(2) M-1 s-1 at I = 40 mM and I = 200 mM, respectively. Amino acid composition and molar absorption coefficients at various wavelengths are reported. Resonances of heme protons and the epsilon H3 group of the ligand methionine of the Fe have been identified in the 1H-NMR spectrum of the reduced as well as the oxidized cytochrome.

Amino Acids↗
PubMed1990

Three-dimensional structure of the mammalian cytoplasmic ribosome.

A three-dimensional reconstruction of the 80 S ribosome from rabbit reticulocytes has been calculated from low-dose electron micrographs of a negatively stained single-particle specimen. At 37 A resolution, the precise orientations of the 40 S and 60 S subunits within the monosome can be discerned. The translational domain centered on the upper portion of the subunit/subunit interface is quite open, allowing considerable space between the subunits for interactions with the non-ribosomal macromolecules involved in protein synthesis. Further, the cytosolic side of the monosome is strikingly more open than the membrane-attachment side, suggesting a greater ease of communication with the cytoplasm, which would facilitate the inwards and outwards diffusion of a number of ligands. Although the 60 S subunit portion of the 80 S structure shows essentially all of the major morphological features identified for the eubacterial 50 S large subunit, it appears to possess a region of additional mass that evidently accounts for the more ellipsoidal form of the eukaryotic subunit.

Animals↗
PubMed1990

The risk approach in diarrheal disease intervention in Grenada.

In a previous paper (Soc. Sci. Med. 23, 995-1002, 1986), we described the development of a method for identifying households at high risk of childhood diarrhea based on a logit analysis of data collected as part of a diarrhea disease control project in Grenada. In this paper, we report the results of a follow-up study designed to replicate the first using data on diarrhea incidence and risk factors collected in a different set of rural communities in Grenada. The findings of the first study were not replicated. There was no significant association between the risk factors and diarrhea in the follow-up study in contrast to the very strong associations found in the first study. As a consequence, it was not possible to construct a logit model from the second data set. The possible reasons for the contrasting sets of results and their implications for the application of the risk approach are discussed with reference to a two by two true-false table.

Diarrhea↗
PubMed1990

Standardized non-invasive assessment of myocardial free fatty acid kinetics by means of 15-(para-iodo-phenyl) pentadecanoic acid (123I-pPPA)scintigraphy: I. Method.

Two simple mathematical functions have been applied for estimating myocardial perfusion and fatty acid metabolism from planar, dynamic 123I-para-phenylpentadecanoic acid (123I-pPPA) studies. The first one uses a modified gamma function and the latter one a biexponential fit. The reciprocal of mean transit time is assumed to represent myocardial blood flow and the fast phase (size and half-time) of the biexponential fit primarily accounts for beta oxidation of 123I-pPPA.

Coronary Circulation↗
PubMed1990

Fuzzy sets-based classification of electron microscopy images of biological macromolecules with an application to ribosomal particles.

Pattern recognition methods based on the theory of fuzzy sets are tested for their ability to classify electron microscopy images of biological specimens. The concept of fuzzy sets was chosen for its ability to represent classes of objects that are vaguely described from the measured data. A number of partitional clustering algorithms and an extensive set of cluster-validity functionals (some already reported and some newly developed) have been applied to a test-data set and to two real-data sets of images. One of the real-data sets corresponded to images of the Escherichia coli 50S ribosomal subunits depleted of proteins L7/L12 and the other set to images of the E. coli 70S monosome in the range of overlap views. These two latter sets had been previously studied by another clustering methodology. The new results obtained by the application of fuzzy clustering techniques will be compared to those previously obtained and some conclusions about the consistency of these classifications will be drawn from this comparison.

Algorithms↗
PubMed1989

The comparative in vitro activity of ofloxacin and selected ophthalmic antimicrobial agents against ocular bacterial isolates.

The in vitro activity of ofloxacin, a new fluoroquinolone anti-infective agent, was evaluated against 419 ocular bacterial isolates of 55 species to determine its potential as a topical agent for the treatment of ocular infections. Other agents tested in this study, in which a modified tube-dilution procedure was used, include norfloxacin, gentamicin, tobramycin, chloramphenicol, and polymyxin B. Ofloxacin demonstrated good to excellent activity against a variety of gram-positive and gram-negative pathogens. The minimum inhibitory concentration against 90% of all bacterial strains tested (MIC90) of ofloxacin was 0.5 microgram/ml for Staphylococcus aureus and S. epidermidis, 2 micrograms/ml for Streptococcus pneumoniae, and 4 micrograms/ml for Pseudomonas aeruginosa. These species were more susceptible to ofloxacin than to any of the nonquinolones tested. The MIC90 of ofloxacin was lower than that of norfloxacin, another quinolone, against S. aureus, S. epidermidis, and St. pneumoniae and equal to that of norfloxacin against P. aeruginosa. Because of its broad spectrum of activity and excellent in vitro activity, we concluded that ofloxacin has the potential for development into a superior topical treatment for ocular infection.

Anti-Bacterial Agents↗
PubMed1989

Three-dimensional reconstruction of mammalian 40 S ribosomal subunit.

The small (40 S) subunit from rabbit reticulocyte ribosomes has been reconstructed from electron micrographs of a negatively stained single-particle specimen to a resolution of 3.85 nm. The reconstruction reveals a morphology consisting of a broad wedge-shaped head structure set atop a quasi-cylindrical body. Distinctive features recognized in two-dimensional projections, such as the beak, back lobes, and feet, can now be localized in three dimensions. By reference to a recent reconstruction of the monomeric 80 S ribosome we can identify the interface and exterior surfaces of the subunit, thus enabling more detailed functional interpretations.

Animals↗
PubMed1989

On the mechanism of inhibition of methanol dehydrogenase by cyclopropane-derived inhibitors.

Extraction of cyclopropanol-inactivated methanol dehydrogenase (MDH) gave a mixture of two interconverting compounds. The same compounds could be prepared from 2,7,9-tricarboxy-1H-pyrrolo[2,3-f]quinoline-4,5-dione (PQQ) and cyclopropanol using a metal oxide (e.g. Ag2O) as a catalyst. Structure elucidation revealed that a C5 3-propanal adduct of PQQ is formed which is present in the extract as a diastereoisomeric mixture of the ring-closed form. Cyclopropanone gave an analogous product, while cyclopropylmethanol behaved as a substrate and was oxidized by the enzyme without ring-opening. From the work described, several arguments can be derived to reject the idea that inactivation proceeds via formation of a pair of free radicals. The mechanism probably consists of a concerted proton abstraction, rearrangement of the cyclopropoxy anion to a ring-opened carbanion and attack of the latter on the electrophilic C5 of PQQ. The measured rate of inactivation (3.7 s-1) is in agreement with such a mechanism. The role of the metal oxide and the enzyme in this process is the catalysis of the addition step and possibly a positioning of the reactants. As only a sole type of quinoprotein alcohol dehydrogenase becomes inhibited, the cyclopropane derivatives studied here can be regarded as mechanism-based inhibitors. The modified PQQ in cyclopropanone-inactivated MDH is fluorescent. A fluorescent intermediate was also observed in the catalytic cycle of MDH with methanol as a substrate. Its rate of formation and decay and the strongly decreased level of fluorescence in the presence of activator are in accordance with the view that the fluorescing species is the previously found oxidized-MDH.substrate (MDHox.S) complex. Since the decomposition of this complex requires activator and model studies have failed so far to mimic the enzyme, it seems that the combination of enzyme and activator is essential for the oxidation of the alcohol substrate.

Alcohol Oxidoreductases↗
PubMed1989

Three-dimensional architecture of the calcium channel/foot structure of sarcoplasmic reticulum.

The calcium channel responsible for the release of Ca2+ from the sarcoplasmic reticulum of skeletal muscle during excitation-contraction coupling has recently been identified and purified. The isolated calcium channel has been identified morphologically with the 'foot' structures which are associated with the junctional face membrane of the terminal cisternae of sarcoplasmic reticulum. In situ, the foot structure extends across the gap of the triad junction from the terminal cisternae of the reticulum to the transverse tubule. We describe here the three-dimensional architecture (3.7 nm resolution) of the calcium channel/foot structure from fast-twitch rabbit skeletal muscle, which we determined from electron micrographs of isolated, non-crystalline structures that had been tilted in the electron microscope. The reconstruction reveals two different faces and an internal structure in which stain accumulates at several interconnected locations, which could empty into the junctional gap of the triad junction. The detailed architecture of the channel complex is relevant to understanding both the physical path followed by calcium ions during excitation-contraction coupling and the association of the terminal cisternae and the transverse tubules in the triad junction.

Animals↗
PubMed1989

Studies on electron transfer from methanol dehydrogenase to cytochrome cL, both purified from Hyphomicrobium X.

Ferricytochrome cL isolated from Hyphomicrobium X is an electron acceptor in assays for homologous methanol dehydrogenase (MDH), albeit a poor one compared with artificial dyes. The intermediates of MDH seen during the reaction are identical with those observed with Wurster's Blue as electron acceptor, indicating that the reaction cycles are similar. The assay showed a pH optimum of approx. 7.0 and scarcely any stimulation by NH4Cl, this being in contrast with assays with artificial dyes, where strong activation by NH4Cl and much higher pH optima have been reported. From the results obtained with stopped-flow as well as steady-state kinetics, combined with the isotope effects found for C2H3OH, it appeared that the dissimilarities between the electron acceptors can be explained from different rate-limiting steps in the reaction cycles. Ferricytochrome cL is an excellent oxidant of the reduced MDH forms at pH 7.0, but the substrate oxidation step is very slow and the activation by NH4Cl is very poor at this pH. At pH 9.0 the reverse situation exists: ferricytochrome cL is a poor oxidant of the reduced forms of MDH at this pH. No C2H3OH isotope effect was observed under these conditions, indicating that substrate oxidation is not rate-limiting, so that activation by NH4Cl cannot be found. Since just the opposite holds for assays with artificial dyes, the poor electron-acceptor capability and the different pH optimum of ferricytochrome cL as well as the insignificant activating effect of NH4Cl (all compared with artificial assays) can be explained. Although different views have been reported on the rate-limiting steps in the systems from Methylophilus methylotrophus and Methylobacterium sp. strain AM1, these are most probably incorrect, as rate-limiting electron transfer between ferrocytochrome cL and horse heart ferricytochrome c can occur. Therefore the conclusions derived for the Hyphomicrobium X system might also apply to the systems from other methylotrophic bacteria. Comparison of the assays performed in vitro (at pH 7.0) having ferricytochrome cL and Wurster's Blue as electron acceptor with methanol oxidation by whole cells shows that the former has similarity whereas the latter has not, this being although ferricytochrome cL is a poor electron acceptor in the assay performed in vitro. The reason for this is the absence of a (natural) activator able to activate the (rate-limiting) substrate oxidation step at physiological pH values.

Alcohol Oxidoreductases↗
PubMed1989

The effect of handedness on spinal and supra-spinal reflex excitability.

Twenty-four subjects, 8 right-handers, 8 left-handers, and 8 familial left-handers, participated in a study aimed at understanding the relations between handedness, cerebral asymmetry, and a possible asymmetry in spinal excitability. This was accomplished by employing 3 different techniques of investigating the H reflex, measuring 3 different parameters of spinal excitability. The recovery curve height (RCH) of left-handers was significantly higher than the RCH of right-handers, and, as well, familial left-handers exhibited higher RCH than non-familial left-handers. The recovery ability was not found to be significantly more inhibited on the dominant side, as would be expected from previous work. Instead, recovery ability was associated with the fractional involvement of the motoneuron pool in the H process (H/M ratio). RCH was highly correlated with both hand preference and hand performance. The above results suggest involvement of a long-loop transcortical reflex in lateral asymmetries.

Adolescent↗
PubMed1989

Image analysis of single macromolecules.

A battery of sophisticated techniques is now available to extract three-dimensional structural information from electron micrographs of biological macromolecules occurring in the form of single particles. One of these techniques, the random-conical reconstruction method, which allows low-dose imaging, has been recently perfected and is being used routinely for the study of ribosomal architecture. The analysis of the 40S mammalian ribosomal subunit serves as an illustration of the various steps of image processing. The use of classification combined with 3-D reconstruction provides the means to investigate variations of the macromolecular structure (deformations, conformational changes, etc.) that are caused by the specimen preparation. An example is provided by the changes in the shape of the 70S monosome of E. coli as it changes its orientation on the carbon grid. The most challenging applications of the techniques discussed are in the area of cryo-microscopy of ice-embedded specimens. First studies of single macromolecules imaged in this way have indicated that the 3-D imaging methods and, specifically, the random-conical reconstruction method, will be applicable under these conditions.

Analysis of Variance↗