Trypanosome U-deletional RNA editing involves guide RNA-directed endonuclease cleavage, terminal U exonuclease, and RNA ligase activities.

We have studied the mechanism of accurate in vitro RNA editing of Trypanosoma brucei ATPase 6 mRNA, using four mRNA-guide RNA (gRNA) pairs that specify deletion of 2, 3, or 4 U residues at editing site 1 and mitochondrial extract. This extract not only catalyzes deletion of the specified number of U residues but also exhibits a novel endonuclease activity that cleaves the input pre-mRNA in a gRNA-directed manner, precisely at the phosphodiester bond predicted in a simple enzymatic model of RNA editing. This cleavage site is inconsistent with a chimera-based editing mechanism. The U residues to be deleted, present at the 3' end of the upstream cleavage product, are then removed evidently by a 3' U-specific exonuclease and not by a reverse reaction of terminal U transferase. RNA ligase can then join the mRNA halves through their newly formed 5' P and 3' OH termini, generating mRNA faithfully edited at the first editing site. This resultant, partially edited mRNA can then undergo accurate, gRNA-directed cleavage at editing site 2, again precisely as predicted by the enzymatic editing model. All of these enzymatic activities cofractionate with the U-deletion activity and may reside in a single complex. The data imply that each round of editing is a four-step process, involving (i) gRNA-directed cleavage of the pre-mRNA at the bond immediately 5' of the region base paired to the gRNA, (ii) U deletion from or U addition to the 3' OH of the upstream mRNA half, (iii) ligation of the mRNA halves, and (iv) formation of additional base pairing between the correctly edited site and the gRNA that directs subsequent nuclease cleavage at the next editing site.

Specific atomic groups and RNA helix geometry in acceptor stem recognition by a tRNA synthetase

Oligonucleotides that recapitulate the acceptor stems of tRNAs are substrates for aminoacylation by many tRNA synthetases in vitro, even though these substrates are missing the anticodon trinucleotides of the genetic code. In the case of tRNAAla a single acceptor stem G⋅U base pair at position 3·70 is essential, based on experiments where the wobble pair has been replaced by alternatives such as I⋅U, G⋅C, and A⋅U, among others. These experiments led to the conclusion that the minor-groove free 2-amino group (of guanosine) of the G⋅U wobble pair is essential for charging. Moreover, alanine-inserting tRNAs (amber suppressors) that replace G⋅U with mismatches such as G⋅A and C⋅A are partially active in vivo and can support growth of an Escherichia coli tRNAAla knockout strain, leading to the hypothesis that a helix irregularity and nucleotide functionalities are important for recognition. Herein we investigate the charging in vitro of oligonucleotide and full-length tRNA substrates that contain mismatches at the position of the G⋅U pair. Although most of these substrates have undetectable activity, G⋅A and C⋅A variants retain some activity, which is, nevertheless, reduced by at least 100-fold. Thus, the in vivo assays are much less sensitive to large changes in aminoacylation kinetic efficiency of 3·70 variants than is the in vitro assay system. Although these functional data do not clarify all of the details, it is now clear that specific atomic groups are substantially more important in determining kinetic efficiency than is a helical distortion. By implication, the activity of mutant tRNAs measured in the in vivo assays appears to be more dependent on factors other than aminoacylation kinetic efficiency.

Building a retrospective collection in pharmacy: a brief history of the literature with some considerations for U.S. health sciences library professionals

This paper argues that historical works in pharmacy are important tools for the clinician as well as the historian. With this as its operative premise, delineating the tripartite aspects of pharmacy as a business enterprise, a science, and a profession provides a conceptual framework for primary and secondary resource collecting. A brief history and guide to those materials most essential to a historical collection in pharmacy follows. Issues such as availability and cost are discussed and summarized in checklist form. In addition, a glossary of important terms is provided as well as a list of all the major U.S. dispensatories and their various editions. This paper is intended to serve as a resource for those interested in collecting historical materials in pharmacy and pharmaco-therapeutics as well as provide a history that gives context to these classics in the field. This should provide a rationale for selective retrospective collection development in pharmacy.

Star scientists and institutional transformation: Patterns of invention and innovation in the formation of the biotechnology industry

The most productive (“star”) bioscientists had intellectual human capital of extraordinary scientific and pecuniary value for some 10–15 years after Cohen and Boyer’s 1973 founding discovery for biotechnology [Cohen, S., Chang, A., Boyer, H. & Helling, R. (1973) Proc. Natl. Acad. Sci. USA 70, 3240–3244]. This extraordinary value was due to the union of still scarce knowledge of the new research techniques and genius and vision to apply them in novel, valuable ways. As in other sciences, star bioscientists were very protective of their techniques, ideas, and discoveries in the early years of the revolution, tending to collaborate more within their own institution, which slowed diffusion to other scientists. Close, bench-level working ties between stars and firm scientists were needed to accomplish commercialization of the breakthroughs. Where and when star scientists were actively producing publications is a key predictor of where and when commercial firms began to use biotechnology. The extent of collaboration by a firm’s scientists with stars is a powerful predictor of its success: for an average firm, 5 articles coauthored by an academic star and the firm’s scientists result in about 5 more products in development, 3.5 more products on the market, and 860 more employees. Articles by stars collaborating with or employed by firms have significantly higher rates of citation than other articles by the same or other stars. The U.S. scientific and economic infrastructure has been particularly effective in fostering and commercializing the bioscientific revolution. These results let us see the process by which scientific breakthroughs become economic growth and consider implications for policy.

Oceanic minerals: Their origin, nature of their environment, and significance

The chemical and isotopic compositions of oceanic biogenic and authigenic minerals contain invaluable information on the evolution of seawater, hence on the history of interaction between tectonics, climate, ocean circulation, and the evolution of life. Important advances and greater understanding of (a) key minor and trace element cycles with various residence times, (b) isotopic sources and sinks and fractionation behaviors, and (c) potential diagenetic problems, as well as developments in high-precision instrumentation, recently have been achieved. These advances provided new compelling evidence that neither gradualism nor uniformitarianism can explain many of the new important discoveries obtained from the chemistry and isotopic compositions of oceanic minerals. Presently, the best-developed geochemical proxies in biogenic carbonates are 18O/16O and Sr/Ca ratios (possibly Mg/Ca) for temperature; 87Sr/86Sr for input sources, Cd/Ca and Ba/Ca ratios for phosphate and alkalinity concentrations, respectively, thus also for ocean circulation; 13C/12C for ocean productivity; B isotopes for seawater pH;, U, Th isotopes, and 14C for dating; and Sr and Mn concentrations for diagenesis. The oceanic authigenic minerals most widely used for chemical paleoceanography are barite, evaporite sulfates, and hydrogenous ferromanganese nodules. Marine barite is an effective alternative monitor of seawater 87Sr/86Sr, especially where carbonates are diagenetically altered or absent. It also provides a high-resolution record of seawater sulfate S isotopes, (evaporite sulfates only carry an episodic record), with new insights on factors affecting the S and C cycles and atmospheric oxygen. High-resolution studies of Sr, Nd, and Pb isotopes of well-dated ferromanganese nodules contain invaluable records on climate driven changes in oceanic circulation.

Biosynthesis of the Monoterpenes Limonene and Carvone in the Fruit of Caraway1 : I. Demonstration of Enzyme Activities and Their Changes with Development

The biosynthesis of the monoterpenes limonene and carvone in the fruit of caraway (Carum carvi L.) proceeds from geranyl diphosphate via a three-step pathway. First, geranyl diphosphate is cyclized to (+)-limonene by a monoterpene synthase. Second, this intermediate is stored in the essential oil ducts without further metabolism or is converted by limonene-6-hydroxylase to (+)-trans-carveol. Third, (+)-trans-carveol is oxidized by a dehydrogenase to (+)-carvone. To investigate the regulation of monoterpene formation in caraway, we measured the time course of limonene and carvone accumulation during fruit development and compared it with monoterpene biosynthesis from [U-14C]Suc and the changes in the activities of the three enzymes. The activities of the enzymes explain the profiles of monoterpene accumulation quite well, with limonene-6-hydroxylase playing a pivotal role in controlling the nature of the end product. In the youngest stages, when limonene-6-hydroxylase is undetectable, only limonene was accumulating in appreciable levels. The appearance of limonene-6-hydroxylase correlates closely with the onset of carvone accumulation. At later stages of fruit development, the activities of all three enzymes declined to low levels. Although this correlates closely with a decrease in monoterpene accumulation, the latter may also be the result of competition with other pathways for substrate.