Incorporation Efficiency of Small Oligo-5'-nucleotide Initiators in the Terminal Deoxyribonucleotide Transferase Reaction<sup>*</sup>

Fn, Hayes; Ve, Mitchell; Rl, Ratliff; Aw, Schwartz; Williams, Dennis

doi:10.1021/bi00875a035

Cited by 34 publications

(19 citation statements)

References 4 publications

(2 reference statements)

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“…The phosphate concentration used is considerably higher than the value (40mM) above which it has been reported [6] that essentially complete inhibition of polymerization on an initiator results. I n our experience [5], concentrations even up to 240 mM yield initiated polymers but at diminishing reaction rates with increasing phosphate concentration. Thermal hyperchromicity measurements (absorbancetemperature profiles) were carried out as in Ratliff et al [20].…”

Section: Methods and N A T E Rmentioning

confidence: 70%

“…The preferred scintillator system was 0.5 ml of sample solution in 10 ml of 6.8 g/1 of 2,5-diphenyloxazole and 0.21 g/l of 2,2'-p-phenylenebis(5-phenyloxazole) in toluene containing 13°/0 by volume of Colosolve Solubilizer CS-3 (Sentol Associates). Polymer length was determined from the isotopic dilution of labeled initiator [5]. Paper disc counting [5] was performed in toluene containing 4 g/1 of 2,5-diphenyloxazole and 0.1 g/l of 2,2'-p-phenylenebis(5-phenyloxazole).…”

Section: Methods and N A T E Rmentioning

confidence: 99%

“…Polymer length was determined from the isotopic dilution of labeled initiator [5]. Paper disc counting [5] was performed in toluene containing 4 g/1 of 2,5-diphenyloxazole and 0.1 g/l of 2,2'-p-phenylenebis(5-phenyloxazole).…”

Section: Methods and N A T E Rmentioning

confidence: 99%

“…According to current knowledge about the chemistry of the cell genome, the arrangement of nucleotide units is a linear mixture of hetero-and homopolymer sequences [l-31. Many synthetic routes are available, all of which may have to be used in combination: organic stepwise linkage [4] and enzymatic chain lengthening [5,6], block linkage [7-111, and replication [12,13].…”

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confidence: 99%

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Synthesis of N‐Acetylated Deoxyribonucleoside 5′‐Triphosphates and their Utilization in Enzymatic Formation of Single‐Stranded Polydeoxyribonucleotides

Hayes¹,

Hansbury²,

Mitchell³

et al. 1968

European Journal of Biochemistry

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The N-acetylated deoxyadenosine and deoxyguanosine 5'4riphosphates (dAAcTP and dGAcTP) have been synthesized and found competent as substrates for enzymatic chain lengthening of polydeoxyribonucleotides without concomitant loss of the acetyl group. The enzyme used was terminal deoxyribonucleotidyltransferase. The addition of dGAc units proceeds without aggregation that self-limits addition of nonacetylated dG units. The resulting polymer 3'-ended with dGAC units is an efficient initiator for subsequent chain lengthening. Hydrogen bonding is not destroyed by N-acetylation of the dA-dT Watson-Crick pair, although it is considerably weakened. As a practical result of this, a polymer 3'-ended with dAAC units is a much more efficient initiator for enzymatic chain lengthening by dT units than is a corresponding polymer with dA units. Venom phosphodiesterase does not hydrolyze a polymer 3'-ended with dAAc U n i t s .Synthesis of polydeoxyribonucleotides with desired sequences can be a key contributor to successful, designed cellular transformation. According to current knowledge about the chemistry of the cell genome, the arrangement of nucleotide units is a linear mixture of hetero-and homopolymer sequences (CBN) ; overlining of a subscript means that the number is the average of a distribution; the superscripts Ac or Ac, following a capital letter show that the nucleotide is acetylated or diacetylated; monomer units in a polymer are referred to by dA, dG*c, etc.; labeling in compounds is sparingly designated, usually just a t the start of an experiment ; the mono-ymole is a micromole of monomer units contained in a polymer. 33.tion of dG units onto an initiator but also very inefficient utilization as an initiator of an oligodeoxyribonucleotide 3'-terminated with a run of dG units. However, when dG units were enzymatically copolymerized with any other nucleotide or combination of nucleotides, aggregation was absent [20]. Aggregation of dC units interferes with use of dC oligomers longer than decamer as initiators for chain lengthening [21]. Hydrogen bonding slows the rate of dT unit addition onto poly dA initiators [22].It has been shown that N-acetylated dG oligomers do not have the property of aggregation [17], and it may readily be inferred that this same chemical modification on other amino-bearing nucleotides could eliminate the other difficulties mentioned above. Therefore, we have synthesized the N-acetylated 5'-triphosphates dGAcTP and dAACTP in order to determine whether these can function as substrates for enzymatic chain lengthening, whether the products retain the N-acetyl group and are devoid of the aggregation and hydrogen-bonding difficulties, and whether the N-acetyl group is Gnally removable with no damage to the polydeoxyribonucleotide. METHODS AND N A T E R

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Section: Methods and N A T E Rmentioning

confidence: 70%

Section: Methods and N A T E Rmentioning

confidence: 99%

Section: Methods and N A T E Rmentioning

confidence: 99%

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confidence: 99%

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Synthesis of N‐Acetylated Deoxyribonucleoside 5′‐Triphosphates and their Utilization in Enzymatic Formation of Single‐Stranded Polydeoxyribonucleotides

Hayes¹,

Hansbury²,

Mitchell³

et al. 1968

European Journal of Biochemistry

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“…[ I] was used [6] to polymerize dTTP onto tritium labeled dT4 [7], resulting in dT, samples with average n values of 110, 170, and 460 as determined by radioisotopic dilution [6] and by agarose column chromatography [8]. Similarly, dATP and dA, gave dAa4,,.…”

Section: Terminal Deoxynucleotidyltransferasementioning

confidence: 99%

Polydeoxythymidylate as template for complementary enzymatic synthesis

et al. 1971

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Polydesoxynucleotide als DNA-Modelle

Wulff

1974

Naturwissenschaften

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Incorporation Efficiency of Small Oligo-5'-nucleotide Initiators in the Terminal Deoxyribonucleotide Transferase Reaction^*

Cited by 34 publications

References 4 publications

Synthesis of N‐Acetylated Deoxyribonucleoside 5′‐Triphosphates and their Utilization in Enzymatic Formation of Single‐Stranded Polydeoxyribonucleotides

Synthesis of N‐Acetylated Deoxyribonucleoside 5′‐Triphosphates and their Utilization in Enzymatic Formation of Single‐Stranded Polydeoxyribonucleotides

Polydeoxythymidylate as template for complementary enzymatic synthesis

Polydesoxynucleotide als DNA-Modelle

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Incorporation Efficiency of Small Oligo-5'-nucleotide Initiators in the Terminal Deoxyribonucleotide Transferase Reaction*

Cited by 34 publications

References 4 publications

Synthesis of N‐Acetylated Deoxyribonucleoside 5′‐Triphosphates and their Utilization in Enzymatic Formation of Single‐Stranded Polydeoxyribonucleotides

Synthesis of N‐Acetylated Deoxyribonucleoside 5′‐Triphosphates and their Utilization in Enzymatic Formation of Single‐Stranded Polydeoxyribonucleotides

Polydeoxythymidylate as template for complementary enzymatic synthesis

Polydesoxynucleotide als DNA-Modelle

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Incorporation Efficiency of Small Oligo-5'-nucleotide Initiators in the Terminal Deoxyribonucleotide Transferase Reaction^*