Biomedical subjects
E Chargaff
Publications and source records attributed to E Chargaff.
A nucleotide phosphotransferase from Escherichia coli. Purification and properties.
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Nicotinamide adenine dinucleotide as substrate of the nucleotide phosphotransferase from Escherichia coli.
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Survey of DNA polymerase activity during the early development of Drosophila melanogaster.
The pattern of DNA polymerase activity in developing Drosophila melanogaster has been studied in seven stages of embryonic development as well as in unfertilized eggs. The crude polymerase-containing extracts, most likely of cytoplasmic origin, utilize, in the following order of decreasing template efficiency, "activated" calfthymus DNA, poly(A).oligo(dT), and poly(A).oligo(U). The highest enzymes levels occur in unfertilized eggs; the activity remains high during the first 9 hr of embryogenesis, but shows a progressive decline in the later stages. Deoxyribonuclease exhibits a similar trend. The unfertilized eggs of two genotypically different females had nearly identical levels of DNA polymerase.
Purification and properties of ribonuclease H of calf thymus.
Ribonuclease H of calf thymus has been purified better than 3000-fold to yield an almost homogeneous preparation. The enzyme, which comprises about 0.03% of the total protein in the initial extract, is a slightly acidic protein (pI = 4.95) of molecular weight of about 64,000, possibly composed of subunits. The enzyme requires a metal ion for activation; the conditions for activation by Mg, Co, and Mn are described. It is inhibited by S-adenosylmethionine. The substrates cleaved were poly(dT).poly(rA) and the DNA-RNA hybrid made from phage f1 DNA; ribosomal RNA was not attacked.
A purine polyribonucleotide synthetase from Escherichia coli.
The isolation, from E. coli B, and partial purification of a purine polyribonucleotide synthetase having several unusual properties is described. The enzyme, which seems to be under strict regulation by several nucleoside triphosphates, requires, after removal of internal primer activity, a primer, such as poly(A), poly(U), or a suitable RNA, but acts without a template. It uses purine ribonucleoside triphosphates as precursors. The uptake of adenylic acid, when ATP is offered alone, is highly stimulated by the presence of GTP, in which case both nucleotides are incorporated into mixed polymers; but GTP as the sole precursor is not utilized. CTP has a strongly inhibitory effect. Other unusual features are the high salt concentration, 0.6 M KCl, at which the enzyme is optimally active and evidence of the existence of a relatively heat-stable protein functioning as an activation factor.
Nucleoside phosphotransferase from carrot. Chemical characterization and investigation of catalytic sites.
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Action of DNA polymerase I of Escherichia coli with DNA-RNA hybrids as templates.
Experiments indicating the ability of the ribo strand of a DNA-RNA template to guide polydeoxynucleotide synthesis by highly purified DNA polymerase I of E. coli (EC 2.7.7.7) are presented. With poly(rA).poly(dT) as template, poly(dT) is formed with a high efficiency, but almost no poly(dA). The specific activity of the enzyme, when tested with this template under suitable conditions, is eight times greater than that found for the poly(dA-dT) template. Single-stranded DNA fractions, with no template activity for DNA polymerase, are converted to efficient templates after their transcription by RNA polymerase. A concerted polymerization reaction, in which the action of DNA polymerase is dependent on that of RNA polymerase, can also be demonstrated with synthetic polydeoxynucleotides and single-stranded fractions of denatured DNA as templates.
DNA polymerase of chicken embryo: purification and properties.
The almost complete purification of the previously reported DNA polymerase of chicken embryo is described; in the final isolation step the enrichment of the enzyme, which comprises about 0.01% of the protein extractable from 10-day-old embryos, is more than 9000-fold. The enzyme is a basic protein (pI = 9.15), of molecular weight about 27,000. It exhibits no nuclease activity, requires Mn(2+) ion for activity, and may contain Zn. It is very sensitive to oxidation.
Mechanism of DNA replication by highly purified DNA polymerase of chicken embryo.
Highly purified DNA polymerase of chicken embryo was used to investigate the mechanism of DNA replication. The templates used included synthetic homopolymer pairs, as well as DNA of bacteriophage f1 and DNA-RNA hybrids prepared by partial or complete transcription of f1 DNA. The evidence suggests that the DNA synthesized is complementary either to the DNA or the RNA strand of the hybrid, depending on the relative lengths of the two strands, with the longer serving as the template and the shorter as the primer. While f1 DNA itself lacks template properties, the f1 DNA-RNA hybrids are efficient templates; and composition studies show the DNA synthesized to be complementary to the DNA strand of the hybrid. These observations suggest a mechanism of DNA replication in which the initial synthesis of a stable primer by RNA polymerase plays a pivotal role.
Preface to a grammar of biology. A hundred years of nucleic acid research.
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On the control of the induction of -galactosidase in synchronous cultures of Escherichia coli.
The mode of induction of beta-galactosidase (EC 3.2.1.23) in synchronously growing cultures of two Hfr strains of Escherichia coli K12 was investigated. Cells can be induced to form the enzyme during any portion of the cell cycle; but when they are grown permanently in the presence of an inducer, enzyme synthesis is discontinuous. The interruption of beta-galactosidase synthesis appears to be geared to the growth cycle: it occurs when the cells are dividing actively. The observation that lac-specific messenger RNA is produced also in the absence of detectable enzyme synthesis suggests the existence of a control mechanism operating on the level of translation.
Nucleic acid polymerases of the developing chicken embryo: a DNA polymerase preferring a hybrid template.
This paper presents a preliminary survey of the nucleic acid polymerases of the developing chicken embryo, especially of the 4-day stage. The predominant activity is that of a DNA polymerase preferring a DNA-RNA hybrid as the template. The enzyme, which is activated by Mn(2+) ions and inhibited by p-chloromercuriphenylsulfonate, copies preferentially the ribo strand of a hybrid, such as poly(rA).poly(dT), but is relatively inactive with all-ribo duplexes. DNA polymerase I of Escherichia coli was also found to use the hybrid template with high efficiency, copying preferentially the ribo strand. With the chicken enzyme, the template activity of denatured DNA was increased tenfold by simultaneous transcription with RNA polymerase. DNA polymerase activity reaches a maximum in the 6- to 8-day chicken embryo and then declines progressively to about one-third of the maximal value in the adult chicken.
Nucleoside phosphotransferase from carrot. Kinetic studies and exploration of active sites.
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[Preface to a grammar of biology. 100 years of nucleic acid research].
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Template properties of complementary fractions of denatured microbial deoxyribonucleic acids.
DNA preparations from seven bacterial species and from E. coli phage T4, and also the complementary L and H fractions into which these DNA specimens, after denaturation, were separated by chromatography on methylated albumin-kieselguhr columns, were studied as templates in the RNA polymerase system, and the nucleotide composition of the RNA products was determined. The RNA transcripts of the separated L and H fractions were found to be faithful copies of the respective DNA fractions. This suggests "transcription analysis" as a sensitive and reliable analytical technique for the determination of the base composition of denatured DNA. The L and H fractions of T4 DNA were shown, both by temperature-absorbance profiles and by transcription analysis, to be mutually complementary. The RNA products formed with intact DNA as the template were not exact copies of the latter; their composition indicated that under our experimental conditions the "heavy" DNA strand is transcribed preferentially.
Complementarity of RNA produced by enzymic transcription of native and denatured B. subtilis DNA.
This paper describes the preparation and some of the properties of the RNA specimens synthesized with the aid of the RNA polymerase of E. coli by transcription of the following DNA templates: (a) undenatured B. subtilis DNA (yielding N-RNA); (b) separated strand fractions L and H isolated by chromatography of the denatured DNA on methylated albumin (yielding L-RNA and H-RNA, respectively). The study of the hybridization behavior of the various RNA products showed that N-RNA, though able to form hybrids with either strand, hybridized with H-DNA to twice as great an extent as with L-DNA. The transcripts of the separated L and H fractions exhibited complete specificity with respect to complexing: L-RNA hybridized only with L-DNA, H-RNA only with H-DNA.
Translational complementarity of RNA transcripts of native and denatured B. subtilis DNA.
Previous evidence has suggested that the L and H fractions into which denatured DNA of B. subtilis can be separated by chromatography are complementary to each other with regard to base compositon, nucleotide sequence, and template properties, and that they may be regarded as families of fragments of the respective DNA strands. The present study tests the proposition that these fractions should also exhibit features of translational complementarity with respect to the DNA-dependent or the RNA-dependent incorporation of amino acids into ribosomes. This has been shown to be the case with the use of two pairs of amino acids: (1) lysine and phenylalanine; (2) proline and glycine.