DNA-RNA hybrid secondary structures

Arnott, Struther; Chandrasekaran, R.; Millane, Rick P.; Park, Hye-Shin

doi:10.1016/s0022-2836(86)80011-0

Cited by 105 publications

(76 citation statements)

References 23 publications

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“…[Deoxynucleotides were used for technical convenience, in particular to avoid byproducts in the synthesis. However, the RNA-DNA hybrid formed with the guide sequence is expected to have the geometry of an RNA A-form helix (24).] The RNA guide sequence was designed to be complementary to both the ACCA of the minihelix and the Phe-bearing aminoacyl phosphate oligonucleotide.…”

Section: Resultsmentioning

confidence: 99%

Peptide synthesis with a template-like RNA guide and aminoacyl phosphate adaptors

Tamura

Schimmel

2003

Proc. Natl. Acad. Sci. U.S.A.

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The genetic code is thought to have developed from an early system of RNA-dependent peptide synthesis. To investigate one kind of template-like peptide synthesis that might emerge from an RNA world, we constructed highly reactive aminoacyl phosphate oligonucleotides as adaptors that bound to RNA guide sequences. The reactive aminoacyl groups mimic a chemistry found in modern protein biosynthesis. Guide sequence interactions with adaptors were borrowed in part from universal contacts seen between tRNAs and rRNA. With these constructions, di-and tripeptides formed in a single guide sequence-dependent reaction. The order of amino acids was not random but directional in a way consistent with substrate reactivities. No ribosomes or ribozymes were required. Thus, aminoacyl phosphate adaptors and RNA guides could, in principle, have been intermediates in the transition from the RNA world to modern template-dependent protein synthesis. D evelopment of modern peptide biosynthesis with RNA templates and aminoacyl-RNA esters in the form of tRNA adaptors was a critical step in the transition from the putative RNA world to the theater of proteins (1). To better understand this development, early work focused on self-condensation reactions with high concentrations of mononucleoside or oligonucleotide aminoacyl esters in reactions that ran for days (2-5).Here we introduced the use of highly reactive aminoacyl phosphate oligonucleotide adaptors and template-like RNA guide sequences. The use of aminoacyl phosphate oligonucleotides was inspired by contemporary systems that use aminoacyl phosphate (mononucleotide) adenylates as intermediates for aminoacyl adaptor (tRNA) synthesis.The free energy of hydrolysis of aminoacyl phosphates is Ϸ3 kcal⅐mol Ϫ1 (1 cal ϭ 4.184 J) greater than that of the aminoacyl esters (aminoacyl-tRNAs) (6) that donate amino acids to growing polypeptide chains in modern template-dependent protein biosynthesis. The aminoacyl esters are formed by transfer of the aminoacyl moieties from aminoacyl phosphates (in the form of aminoacyl adenylates) to the 3Ј ends of adaptors (tRNAs) in reactions catalyzed by aminoacyl-tRNA synthetases (7-9). With this transfer reaction in mind, we envisioned a scenario where the ester was eliminated and the high energy of the aminoacyl phosphate was captured by the adaptor itself. For this purpose, we designed a system of three components (Fig. 1). These were an aminoacyl minihelix, an aminoacyl phosphate oligonucleotide, and an RNA guide sequence. The idea was that, with the proper design of each of these three components, peptide synthesis might occur simply on mixing them together. Materials and MethodsSynthesis and Preparation of Substrates. Minihelix Ala and the template oligoribonucleotides were synthesized on an Expedite 8909 synthesizer (PE Biosystems). Oligodeoxyribonucleotides with a phosphate group at the 5Ј position were synthesized by Invitrogen. Minihelix Ala and the template oligoribonucleotides were deprotected by using the methods of Wincott et al. (10). Escheric...

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Section: Resultsmentioning

confidence: 99%

Peptide synthesis with a template-like RNA guide and aminoacyl phosphate adaptors

Tamura

Schimmel

2003

Proc. Natl. Acad. Sci. U.S.A.

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“…Initially, the 18-nt duplex region of the tRNA-viral RNA complex adopts an A-form helical geometry, but after incorporation of dNTPs, the primer-template complex becomes a DNA-RNA hybrid and assumes an intermediate conformation in solution (12)(13)(14). X-ray crystallographic structures of RT bound to primer-template complexes (D18 or RNA polypurine tract (PPT) primer annealed to a complementary DNA template) reveal a bend in these complexes of about 40° (15)(16)(17).…”

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confidence: 99%

Efficient Initiation of HIV-1 Reverse Transcriptionin Vitro

Iwatani¹,

Rosen²,

Guo³

et al. 2003

Journal of Biological Chemistry

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“…Lane 1 shows the reactivity of the polyclonal antiserum. The fuzzy bands in the molecular mass region 2). This protein, however, did not fractionate with RNase H activity and it is immunologically unrelated to the 160-kDa yeast reverse transcriptase [6,81.…”

Section: Characterization Of the Rut Anti-(yeast Rnase H(55)]mentioning

confidence: 87%

“…These nucleases are thought to recognize specifically RNA/DNA hybrid molecules that appear to have an A-like conformation or a heteromerous structure, in which the two strands of the helix adopt different conformations [2]. RNA/ DNA hybrids occur only transiently during transcription and replication, as well as during reverse transcription of RNA to DNA, and hence resemble potential regulatory sites of negative control by RNase H. In Escherichia coli, RNase H appears to play a crucial role in the regulation of the switch between alternative pathways for the initiation of DNA replication [3], reviewed in [4].…”

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Identification of a yeast ribonuclease H as an Sm antigen

Karwan

Kindås‐Mügge

1989

European Journal of Biochemistry

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We have isolated a 55-kDa enzyme from Saccharomyces cerevisiue on the basis of its ability to hydrolyze specifically the RNA moiety of RNA/DNA hybrids [RNase H(55)]. Remarkably, monospecific anti- [RN ase H(55)] antibodies revealed that the protein associates with several small RNAs, including some of the essential yeast spliceosomal snRNAs. Moreover, immunoprecipitation as well as immunoblotting experiments demonstrated that the yeast enzyme reacts (a) with human anti-Sm autoantisera, (b) with a monoclonal antibody specific for the human snRNP proteins B/B', but (c) not with U1-ribonucleoprotein-specific autoantibodies. These rcsults disclosed a hitherto unexpected degree of evolutionary conservation in snRNP protein structure between yeast and man. Additionally, our findings suggested a re-evaluation of the enzymatic mechanism of RNases H which recognize both RNA and RNA/DNA hybrids.Enzymes that hydrolyze the RNA strand of RNA/DNA hybrids (RNases H) have been described from various cell types [l]. These nucleases are thought to recognize specifically RNA/DNA hybrid molecules that appear to have an A-like conformation or a heteromerous structure, in which the two strands of the helix adopt different conformations [2]. RNA/ DNA hybrids occur only transiently during transcription and replication, as well as during reverse transcription of RNA to DNA, and hence resemble potential regulatory sites of negative control by RNase H. In Escherichia coli, RNase H appears to play a crucial role in the regulation of the switch between alternative pathways for the initiation of DNA replication [3], reviewed in [4]. In contrast, the biological role of eukaryotic RNases H has remained elusive [l].We set out to study RNase H function in Saccharomyces cerevisiae because of the greater possibilities for performing reversed genetics in this organism. Recently, we isolated and partly characterized a ribonuclease H with a molecular mass of 70 kDa from a protease-deficient yeast strain [RNase H(70) The recent finding that monospecific anti-[yeast RNase H(55)] antibodies cross-reacted with a human protein from HeLa cells (Karwan, R., unpublished results) prompted us to examine the potential interactions of RNase H(55) with higher-order cellular structures.We describe here the surprising finding of the association of RNase H(55) with several small RNAs. Besides tRNA, we

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DNA-RNA hybrid secondary structures

Cited by 105 publications

References 23 publications

Peptide synthesis with a template-like RNA guide and aminoacyl phosphate adaptors

Peptide synthesis with a template-like RNA guide and aminoacyl phosphate adaptors

Efficient Initiation of HIV-1 Reverse Transcriptionin Vitro

Identification of a yeast ribonuclease H as an Sm antigen

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