Studies on polypeptide-chain-elongation factors from an extreme thermophile, Thermus thermophilus HB8. 3. Molecular properties.

Molecular properties of the polypeptide chain elongation factors from Thermus thermophilus HB8 have been investigated and compared with those from Escherichia coli. 1. As expected, the factors purified from T. thermophilus were exceedingly heat-stable. Even free EF-Tu not complexed with GDP was stable after heating for 5 min at 60 degrees C. 2. GDP binding activity of T. thermophilus EF-Tu was also stable in various protein denaturants, such as 5.5 M urea, 1.5 M guanidine-HCl, and 4 M LiCl. 3. Amino acid compositions of EF-Tu and EF-G from T. thermophilus were similar to those from E. coli. On the other hand, amino acid composition of T. thermophilus EF-Ts was considerably different from that of E. coli EF-Ts. 4. In contrast to E. coli EF-Tu, T. thermophilus EF-Tu contained no free sulfhydryl group, but one disulfide bond. The disulfide bond was cleaved by sodium borohydride or sodium sulfite under native conditions. The heat stability of the reduced EF-Tu . GDP, as measured by GDP binding activity, did not differ from that of the untreated EF-Tu . GDP. 5. T. thermophilus EF-Ts contained, in addition to one disulfide bond, a sulfhydryl group which could be titrated only after complete denaturation of the protein. 6. Under native conditions one sulfhydryl group of T. thermophilus EF-G was titrated with p-chloromercuribenzoate, while the rate of reaction was very sluggish. The sulfhydryl group appears to be essential for interaction with ribosomes, whereas the ability to form a binary GDP . EF-G complex was not affected by its modification. The protein contained also one disulfide bond. 7. Circular dichroic spectra of EF-Tu from T. thermophilus and E. coli were very similar. Binding of GDP or GTP caused a similar spectral change in both. T. thermophilus and E. coli EF-Tu. On the other hand, the spectra of T. thermophilus EF-G and E. coli EF-G were significantly different, the content of ordered structure being higher in the former as compared to the latter.     

Purification of tubulin from bovine brain and its interaction with guanine nucleotides.

A rapid and sensitive assay for [3H]GTP binding activity of tubulin has been developed. This assay method is based on the quantitative retention of [3H]GTP. Tubulin complex on a nitrocellulose membrane filter. It was also found that bovine brain tubulin is markedly stablized by glycerol and GTP against denaturation. A large-scale purification of bovine brain tubulin was achieved using the new assay procedure and by the inclusion of glycerol and GTP in a buffer solution used for column chromatograph. The purified tubulin could be stored at -80degrees in the presence of glycerol and GTP for at least a year without any apprecialbe loss of [3H]GTP- and [3H]colchicine binding activities. The interaction of tubulin with guanine nucleotides was also studied using the nitorcellulose membrane filter procedure. It was found that the binding of [3H]GTP to tubulin with an empty exchangeable site proceeded promptly within k sec while the exchange of [3H]GTP- with a GTP-tubulin complex in which the exchangeable site had been occupied with unlabeled GTP occured more slowly. The dissociation constants for GTP and GDP at the exchangeable site of tubulin were determined as 0.5 times 10-6M and 1.9 times 10-6M, respectively. 5'-Guanylylimidodiphosphate could interact, although less strongly, with tubulin at this site, whereas the interaction of other nucleoside triphosphates includint ATP, CTP, UTP, and 5'-guanylyl methylenediphosphonate was very weak, if it occured at all. The presence of Mg2+ and a free sulfhydryl group was found to be essential for binding of [3H]GTP to tubulin. Ca2+ was found to replace Mg2+ in this binding reaction.     

River to river: First evidence of eel movement between distant rivers via the sea

Environmental Biology of Fishes - Eel movement patterns have been frequently studied to learn about their movements within the fresh- and brackish waters of the same river before their spawning...     

Late Quaternary Environmental and Human Impacts on the Mitochondrial DNA Diversity of Four Commensal Rodents in Myanmar

Journal of Mammalian Evolution - We addressed the spatiotemporal characteristics of four commensal rodent species occurring in Myanmar in comparison with other areas of the Indo-Malayan region. We...     

Rapid Severing and Motility of Chloroplast-Actin Filaments Are Required for the Chloroplast Avoidance Response in Arabidopsis

Phototropins (phot1 and phot2 in Arabidopsis thaliana) relay blue light intensity information to the chloroplasts, which move toward weak light (the accumulation response) and away from strong light (the avoidance response). Chloroplast-actin (cp-actin) filaments are vital for mediating these chloroplast photorelocation movements. In this report, we examine in detail the cp-actin filament dynamics by which the chloroplast avoidance response is regulated. Although stochastic dynamics of cortical actin fragments are observed on the chloroplasts, the basic mechanisms underlying the disappearance (including severing and turnover) of the cp-actin filaments are regulated differently from those of cortical actin filaments. phot2 plays a pivotal role in the strong blue light–induced severing and random motility of cp-actin filaments, processes that are therefore essential for asymmetric cp-actin formation for the avoidance response. In addition, phot2 functions in the bundling of cp-actin filaments that is induced by dark incubation. By contrast, the function of phot1 is dispensable for these responses. Our findings suggest that phot2 is the primary photoreceptor involved in the rapid reorganization of cp-actin filaments that allows chloroplasts to change direction rapidly and control the velocity of the avoidance movement according to the light’s intensity and position.     

Synthesis of 2′,3′-Dideoxypurinenucleosides via the Palladium Catalyzed Reduction of 9-(2,5-Di-O-acetyl-3-bromo-3-deoxy-β-d-xylofuranosyl)purine Derivatives

Abstract

Practical method to produce 2′,3′-dideoxypurinenucleosides from 9-(2,5-di-O-acetyl-3-bromo-3-deoxy-β-D-xylofuranosyl)purines (1) was developed. High ratio of 2′,3′-dideoxynucleoside to 3′-deoxyribonucleoside was obtained by selecting the reaction conditions (solvent, pH and/or base), or changing 2′-acyloxy leaving group. The reaction mechanism was studied by deuteration experiments of 1a and 1-(3,5-di-O-acety1-2-bromo-2-deoxy-β-D-ribofuranosyl)thymine (12).