Interactions of yeast tRNAPhe with ribosomes from yeast and Escherichia coli. A fluorescence spectroscopic study.

The interaction of ethidium-labeled tRNAPhe from yeast with ribosomes from yeast and Escherichia coli was studied by stead-state measurements of fluorescence intensity and polarization. The ethidium label was covalently inserted into either the anticodon or the dihydrouridine loop of the tRNA. The codon-independent formation of a tRNA-ribosome complex led to only a moderate increase of the observed fluorescence polarization indicating a considerable internal mobility of the labeled parts of the tRNA molecule in the ribosome complex. When the ribosome complex was formed in the presence of poly(U), the probes both in the dihydrouridine loop and in the anticodon loop were strongly immobilized, the latter exhibiting a substantial increase in fluorescence intensity. A smaller intensity change was observed when E. coli ribosomes were used, although the extent of immobilization was found to be similar in this case. Competition experiments with non-labeled tRNAPhe showed that the labeled tRNAPheEtd was readily released from the complex with yeast ribosomes when poly(U) was absent, whereas in the presence of poly(U) it was bound practically irreversibly. The finding that the mobility of a probe in the dihydrouridine loop is affected by the codon-anticodon interaction on the ribosome suggests a conformational change of the ribosome-bound tRNA which may involve opening of the tertiary structure interactions between the dihydrouridine and the TpsiC loop.     

Nuclear magnetic resonance and fluorescence studies of substrate-induced conformational changes of histidine-binding protein J of Salmonella typhimurium.

The histidine-binding protein J of Salmonella typhimurium binds L-histidine as a first step in the high-affinity active transport of this amino acid across the cytoplasmic membrane. High-resolution nuclear magnetic resonance spectroscopy has been used to monitor the conformation of histidine-binding protein J in the presence and absence of substrate. Evidence is presented to show that this binding protein undergoes a conformational change involving a substantial number of amino-acid residues (including tryptophans) in the presence of L-histidine and that this change is specific for L-histidine. In order to monitor the involvement of tryptophan residues in the substrate-induced conformational change, 5-fluorotryptophan has been incorporated biosynthetically into the histidine-binding protein J using a tryptophan autotroph of Salmonella typhimurium. There are no significant differences in the conformation and binding activity between the 5-fluorotryptophan-labeled and the normal histidine-binding protein J. Proton and fluorine-19 nuclear magnetic resonance studies of the 5-fluorotryptophan-labeled binding protein show that at least one (and possibly two) of the tryptophan residues undergo(es) a change toward a more hydrophobic environment in the presence of L-histidine. These observations are supported by fluorescence data and by differences in the reactivity of the tryptophan residues of this protein toward N-bromosuccinimide in the presence and absence of substrate. The present results are consistent with models for the action of periplasmic-binding proteins in shock-sensitive transport systems of gram-negative bacteria which require a substrate-induced conformational change prior to the energy-dependent translocation of substrates.     

Computer analysis of Staphylococcus aureus phage typing data from 1957 to 1975, citing epidemiological trends and natural evolution within the phage typing system

Computer analysis of Staphylococcus aureus phage ty ping data collected for over 18 years in a large research hospital showed a drastic decrease in the number of hospital epidemic strains. Phage lysis patterns gradually modified from those of earlier years and were a reflection of changes within the S. aureus reservoir, and not within the typing phages, since the typing phages were used from stable lyophilized stocks. There was increasing cross-lysis of S. aureus strains by phages of lytic groups I, II, and III, such that this grouping was no longer epidemiologically valid. A 61% increase in unique strains occurred from the period 1957 to 1975. Disappearance of the widely recognized epidemic strains was followed by a proliferation of unique strains with individual phage patterns. These increased from 38% in the period 1957 to 1962 to 62% in the period 1969 to 1975, indicating a trend toward a "one patient-one strain" situation. Nontypable strains decreased in more recent years from 16% (1957 to 1975) to 7% in 1978, following introduction of phages 94, 96, 292, and D-11. Pandemic S. aureus strain 80/81 first appeared in this hospital in 1959, 5 years after it was first reported in the United States. Strain 80/81 disappeared from the hospital in 1963, partly due to the advent of methicillin.     

Metabolic basis of differential susceptibility of two forest lepidopterans to diflubenzuron

Sixth instar Choristoneura occidentalis Freeman and 5th instar Orgyia pseudotsugata (McDunnough) were fed ring-labelled ¹⁴C-diflubenzuron coated on Douglas-fir needles. After ingestion the gut was purged of radioactive needle residues with artificial diet. Radioactivity in body and frass extracts and residues were measured. Diflubenzuron in extracts were separated from metabolites by thin layer chromatography.Ninety percent and 86% of the ingested diflubenzuron passed through the body without being absorbed in C. occidentalis and O. pseudotsugata, respectively. Bodies of C. occidentalis retained 0.3% of ingested diflubenzuron, while O. pseudotsugata retained 5.3%. This difference in % diflubenzuron retention in the body was negatively correlated with % relative metabolism in the two species. Relative toxicities of diflubenzuron to the two species may be related to % retained diflubenzuron which may be due to a significant difference in relative metabolism.

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Zusammenfassung Sechstraupen von Choristoneura occidentalis Freeman und Fünftraupen von Orgyia pseudotsugata (McDunnough) wurden Douglasiennadeln gefüttert, die mit ringmarkiertem ¹⁴C-Diflubenzuron überzogen waren. Nach der Nahrungsaufnahme wurde der Darm von radioaktiven Nadelresten befreit durch Fütterung einer künstlichen Diät. Die Radioaktivität der Körper- und der Kotextrakte und der Nahrungsreste wurde gemessen. In den Extrakten wurde durch Dünnschichtchromatographie Diflubenzuron von den Metaboliten getrennt.90% resp. 86% des aufgenommenen Diflubenzuron ging durch den Körper hindurch, ohne von den Raupen von C. occidentalis und O. pseudotsugata aufgenommen zu werden. Der Körper von C. occidentalis behielt 0.3%, derjenige von O. pseudotsugata 5.3% des aufgenommenen Diflubenzurons zurück. Dieser Unterschied im prozentualen Zurückhalten von Diflubenzuron war negativ korreliert mit dem prozentualen Metabolismus in beiden Arten. Die relative Giftigkeit von Diflubenzuron auf beide Arten mag abhängig sein vom Prozentsatz zurückgehaltenen Wirkstoffs. Dieser dürfte abhängig sein vom wesentlich unterschiedlichen Ausmass des Metabolismus.

     

Does copper-d-penicillamine catalyze the dismutation of O2−?

It has been reported (M. Younes and U. Weser, 1977, Biochem. Biophys. Res. Commun.78, 1247–1253; E. Lengfelder and E. F. Elstner, 1978, Hoppe-Seyler's Z. Physiol. Chem.359, 751–757) that the complex [Cu(I)₈Cu(II)₆(D-penicillamine)₁₂Cl]^5?-efficiently catalyzes the dismutation of O₂^? and that this activity is resistant to both EDTA and CN^?. However, careful study has demonstrated that this complex is unable to catalyze the dismutation of O₂^?, but that it slowly decomposes to simpler copper complexes which are active. Moreover, the activity which is observed is suppressed by EDTA or by Chelex 100 treatment.     

Calcium-activated thiol-proteinase activity in the fusion of rat erythrocytes induced by benzyl alcohol.

1. Rat erythrocytes were fused by incubation with benzyl alcohol and Ca2+. 2. Cell fusion was inhibited by EGTA, N-ethylmaleimide, tetrathionate, iodoacetamide, cystamine, Tos-Lys-CH2Cl, and to a lesser extent by Tos-Phe-CH2Cl. Phenylmethanesulphonyl fluoride, Tos-Arg-OMe and histamine did not inhibit cell fusion. 3. Gel electrophoresis of membrane proteins from "ghosts" of the erythrocytes treated with benzyl alcohol showed that a high-molecular-weight polymer was present: this was consistent with the entry into the cells of Ca2+ and the activation of a transglutaminase enzyme. 4. In the treated cells the proteins corresponding to bands 2 and 3 in human erythrocytes were decreased, and a polypeptide with a slightly greater mobility than band 3 was produced. 5. These changes were inhibited by EGTA, N-ethylmaleimide, tetrathionate, iodoacetamide, cystamine, and Tos-Lys-CH2Cl, but not by phenylmethanesulphonyl fluoride, Tos-Arg-OMe, or histamine. 6. The intramembraneous particles of the P-fracture face of cells treated with benzyl alcohol to induce fusion were decreased in number and were susceptible to cold-induced aggregation; both of these phenomena were markedly inhibited to EGTA, and partially inhibited by Tos-Lys-CH2Cl and N-ethylmaleimide. 7. These several observations indicate that a Ca2+-activated thiol-proteinase, which acts to degrade membrane proteins and to give freedom of lateral movement to intramembranous particles, may be essential feature of membrane fusion in this system. 8. It is suggested that this proteinase may act to degrade spectrin-binding proteins that attach band-3 protein to the erythrocyte cytoskeleton.     

Competitive interactions and the availability of sleeping sites for a diurnal coral reef fish

A field study was made to test whether the population size of a diurnal reef fish, the wrasse Thalassoma bifasciatum (Bloch), was limited by inter- or intraspecific competition for sleeping shelter. T. bifasciatum is often attacked at dusk by two small territorial damselfishes, Eupomacentrus dorsopunicans (Poey) and E. planifrons (Cuvier). Although these three species sleep in the same general habitats, there are qualitative differences in the types of holes they use and how they use them. Wrasse holes are usually in these damselfishes' territories, but damselfish attacks do not prevent wrasses entering holes. Wrasses infrequently defend their holes intraspecifically. They regularly change their holes, with little intra- or interspecific aggressive interaction. When its hole is removed, a wrasse is late in retiring but finds a hole near its old one with little aggressive interaction, and does not have a higher mortality rate. Empty wrasse holes are rarely refilled, and then only by conspecifics. Wrasses added to reefs find unoccupied holes and do not usurp other fishes' holes. Damselfish defend their eggs and food against the wrasse, but not their sleeping shelter, nor living space per se. Sleeping sites are not limiting the wrasse, but are present in a surplus. Intraspecific hole defense by a wrasse prevents a delay in its retiring that would increase the risk of crepuscular predation on it.     

Influence of molecular configuration on the passage of macromolecules across the glomerular capillary wall

The influence of molecular configuration on the filtration of macromolecules across glomerular capillary walls was examined by comparing fractional clearances of two uncharged polysaccharides of distinctly different molecular configuration in the Munich-Wistar rat. The macromolecules employed were dextran, a slightly branched polymer of glucopyranose, and ficoll, a highly cross-linked copolymer of sucrose and epichlorohydrin. Differences in effective shape between these two polymers were determined from measurements of several physical properties of aqueous solutions containing either dextran or ficoll. It was found that dextran is best represented as a prolate ellipsoid with axial ratios of 4, 9, and 16 for molecules with Stokes-Einstein radii of 22, 32, and 40 A, respectively. On the other hand, ficoll is more closely approximated as spherical since the axial ratio was found to be between 1 and 2 for all molecular sizes. Fractional clearances of dextran and ficoll ranging in effective radius from 18 to 44 A were determined in each of seven Munich-Wistar rats. Fractional clearances of dextran were found to be greater than those of ficoll, the difference being significant for molecular radii ranging from 24 to 44 A. In addition, as shown previously for dextran, ficoll was found to be neither secreted nor reabsorbed by the renal tubules. These results, therefore, suggest that in addition to molecular size and charge, molecular configuration is also a determinant of the filtration of macromolecules across the glomerular capillary wall.     

Interactions of Folch-Lees proteolipid apoprotein with planar lipid bilayers

Summary Water-soluble Folch-Lees proteolipid apoprotein from bovine CNS white matter induces a voltage-dependent conductance in black lipid membranes. Na⁺ is required for the induced conductance change but the established conductance has very low ionic selectivity. The induced conductance fluctuates with a minimum amplitude of 10^–11–10^–10 mho. The magnitude of the conductivity change is dependent on protein concentration and on the composition of lipid bilayers. At a fixed voltage the induced conductance of a phosphatidylcholine-cholesterol membrane is proportional to the sixth power of the protein concentration and the first power of Na⁺ concentration. The interactions between the apoprotein and the lipids are both electrostatic and hydrophobic, but the interaction leading to the conductance increase appears to be mainly hydrophobic. Both the increase in conductance and the current fluctuations remain after extensive washing of the chambers to remove the protein. Furthermore, pronase or glutaraldehyde added to either the cis or trans side of the membrane does not affect the apoprotein-established conductance. However, if the bilayer is formed in the presence of both the apoprotein and pronase or if the apoprotein is treated with pronase prior to its addition to the chamber, no conductance change is observed. The association of the apoprotein with the membrane thus appears to render the protein inaccessible to proteolytic digestion, suggesting that the apoprotein is at least partially imbedded in the membrane interior.