Nucleotide sequences of the separate origins of synthesis of bacteriophage G4 viral and complementary DNA strands.

Fiddes, John C.; Barrell, Bart; Godson, G. N.

doi:10.1073/pnas.75.3.1081

Cited by 84 publications

(29 citation statements)

References 30 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The G4ori c has been extensively studied as a model origin of DNA replication. Deletion studies have shown that the G4ori c region is a 140-nt, noncoding sequence containing potential secondary structure: stem-loops I, II, and III (5). Primase synthesizes pRNA at a 5Ј CTG 3Ј site, close to the 3Ј side of stem-loop I (7,8,18).…”

mentioning

confidence: 99%

Interaction of Escherichia coli primase with a phage G4ori(c)-E. coli SSB complex

Sun

Godson

1996

J Bacteriol

Self Cite

View full text Add to dashboard Cite

We earlier reported that Escherichia coli single-stranded DNA-binding protein (SSB) bound in a fixed position to the stem-loop structure of the origin of complementary DNA strand synthesis in phage G4 (G4ori c ), leaving stem-loop I and the adjacent 5 CTG 3, the primer RNA initiation site, as an SSB-free region (W. Sun and G. N. Godson Primase is required to initiate DNA replication in prokaryotes and eucaryotes. It functions to synthesize short primer RNAs (pRNA) on the separated strands of the replication fork, which provide primed templates with free 3Ј-OH terminus for DNA polymerase to elongate and synthesize the complementary DNA strand (4,13,14,25). Several prokaryotic priming systems have been established to study Escherichia coli primase (referred to as primase in this paper) function in vitro, such as cloned E. coli chromosomal origin of DNA replication (oriC) (9, 24), the origin of complementary DNA strand synthesis (ori c ) of single-stranded (ss) E. coli phages (e.g., X174[1]; G4 [3,25]; and st-1, ␣3, and K [2, 17]), double-stranded phage (e.g., [26]), and E. coli plasmid origins of DNA replication (e.g., R1 [11]). The simplest system is ss phage ori c exemplified by phage G4. In this system, primase in association with E. coli ss DNA-binding protein (SSB) synthesizes 29 nucleotides (nt) of pRNA from a unique 5Ј CTG 3Ј sequence in a region of the viral DNA strand called G4ori c (10). This is in contrast to other priming systems in which primase acts in concert with several other initiation proteins and synthesizes pRNA at many sites on the template DNA, although almost always initiating at 5Ј CTG 3Ј sequence (27). The G4ori c has been extensively studied as a model origin of DNA replication. Deletion studies have shown that the G4ori c region is a 140-nt, noncoding sequence containing potential secondary structure: stem-loops I, II, and III (5). Primase synthesizes pRNA at a 5Ј CTG 3Ј site, close to the 3Ј side of stem-loop I (7,8,18). Initiation of DNA replication in G4-related ss phages ␣3, st-1, and K (17) is similar to that of G4ori c .Several early studies provided information on the interaction of primase with the ori c DNA and SSB. A nuclease footprinting study on the interaction of primase with SSB-coated Kori c demonstrated that regions of the stem-loop structure were involved in primase binding (16). Then a stoichiometric study using gel filtration reported that two primase molecules bound on one G4ori c that was cloned into phage M13 ss-DNA (19). In a more recent study (21) to dissect the interactions of SSB and primase with G4ori c , we have demonstrated that two SSB tetramers bound on G4ori c in a fixed position. This resulted in the formation of a unique structure with two SSB tetramers (i.e., an SSB octamer) bound to the stem-loop region and other SSB tetramers bound on the 5Ј and 3Ј sides, leaving approximately 30 nt of free uncoated ss-DNA between the central SSB octamer and the adjacent SSB tetramers (or octamers). The 5Ј CTG 3Ј pRNA initiation site was located in the SSB-free linke...

show abstract

mentioning

confidence: 99%

Interaction of Escherichia coli primase with a phage G4ori(c)-E. coli SSB complex

Sun

Godson

1996

J Bacteriol

Self Cite

View full text Add to dashboard Cite

show abstract

“…Based on the identification offour repeated sequences which lie 24-63 nt upstream from the termination sites of D-loop strands, Doda et aL (1) postulated that the arrest of D-loop strand elongation is a template sequenceassociated event. The vector M13Goril contains the bacteriophage G4 cDNA strand origin (G4O,) (2) which has the potential to form two stable hairpin structures.…”

mentioning

confidence: 99%

Template-directed pausing in in vitro DNA synthesis by DNA polymerase a from Drosophila melanogaster embryos.

Kaguni¹,

Clayton²

1982

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

View full text Add to dashboard Cite

The activity of Drosophila melanogaster DNA polymerase a on DNA-primed single-stranded DNA templates has been examined. The We have examined the influence of template sequence and structure on polymerization by Drosophila melanogaster DNA polymerase a by constructing single-stranded templates for in vitro DNA synthesis which contain DNA sequences of functional importance in vivo. The heavy-strand (H-strand) origin region of mouse mtDNA was inserted into the duplex replicative form ofthe single-stranded bacteriophage vector Ml3Coril. The 3' ends of the five major displacement-loop (D-loop) strands ofmouse mtDNA have been mapped to clusters ofthree to five nucleotides (nt) on the light-strand (L-strand) template in the D-loop region (1). Based on the identification offour repeated sequences which lie 24-63 nt upstream from the termination sites of D-loop strands, Doda et aL (1) postulated that the arrest of D-loop strand elongation is a template sequenceassociated event. The vector M13Goril contains the bacteriophage G4 cDNA strand origin (G4O,) (2) which has the potential to form two stable hairpin structures.Sites of interruption of elongation on single-stranded DNA by Escherichia coli DNA polymerase II (3), vaccinia DNA polymerase (4), and T4 DNA polymerase (5) have been correlated with the positions of predicted stable hairpin structures in the DNA template. Furthermore, the affinity ofKB cell polymerase a for primed single-stranded homo-and heteropolymers was shown to be markedly affected by the base composition of the primer template and by added unprimed inhibitor DNAs (6).We (9), and E. coli single-strand binding protein (SSB) (fraction IV, 4 x 104 units/mg) (10) were kindly provided by B. Sauer, P. Burgers, S. Scherer, and J. Kaguni, respectively. M13-type single-stranded DNAs were prepared as described (11). Primer-templates were prepared by hybridizing purified restriction fragments (at twice the molar concentration of single-stranded circles) to M13Hori8 and M13Hori3 singlestranded circles (15 ,.g/ml) for 5 hr at 370C in hybridization buffer (0.6 M NaCV0.2 M Tris HCl, pH 7.5/0.02 M EDTA/ 50% formamide) after denaturation ofthe restriction fragments.Construction and Isolation of M13Hori8 and M13Hori3. The mouse mtDNA heavy-strand (H-strand) origin is contained within a 1471-base-pair (bp) HincII/Bal I DNA fragment extending from position 15,188 to position 364 on the mouse mtDNA sequence (12). The vector M13Goril (13)

show abstract

“…These include the use of closed circular DNA (11), synthetic poly(dAT) *poly(dAT) (12), or intracellular circular DNA (13) (29)(30)(31)(32). Hairpin loops of 44 and 60 base pairs were isolated from M13 virus DNA (30); similar helical segments have been identified and sequenced in phage G4 viral DNA (31). These structures were isolated first as cores of single-stranded DNA that resist digestion by S 1 nuclease or other enzymes specific for unpaired nucleic acids.…”

Section: Discussionmentioning

confidence: 99%

Secondary Structure in Denatured DNA is Responsible for Its Reaction with Antinative DNA Antibodies of Systemic Lupus Erythematosus Sera

Stollar¹,

Papalian²

1980

J. Clin. Invest.

View full text Add to dashboard Cite

A B S T R A C T Experiments were designed to determine the basis for the strong competitive reaction of denatured DNA with systemic lupus erythematosus (SLE) antinative DNA antibodies. Secondary structure in denatured DNA was reflected in hyperchromicity upon heating and in multiphase kinetics ofits digestion by Si nuclease. Partial digestion by Si nuclease completely eliminated the ability of denatured DNA to react with antidenatured DNA antibodies, but not its ability to react with SLE sera. Sl nuclease-resistant cores were isolated from extensively digested denatured DNA. These cores had secondary structure, including some stable fold-back helical regions. The cores, from 20 to several hundred base pairs in size, competed with native DNA for binding by SLE sera. Other experiments measured reactions of denatured DNA under conditions that affected its secondary structure content. Its competitive activity decreased as temperature was increased from 00 to 37°C, whereas the activity of native DNA was not altered in this temperature range. With DNA pieces of 90-110 base pairs, native fragments were much more effective than the denatured fragments, in which stable helical structure is less likely to occur than in high molecular weight denatured DNA. Competitive assays with mononucleotides, oligonucleotides, homopolymers, and RNA-DNA hybrids also indicated that two strands of polydeoxyribonucleotide were required for optimal reactions with these SLE serum antibodies. The antibodies can measure stable helical regions in denatured DNA; they may also stabilize short helical regions that occur in an equilibrium of conformational forms.

show abstract

Nucleotide sequences of the separate origins of synthesis of bacteriophage G4 viral and complementary DNA strands.

Cited by 84 publications

References 30 publications

Interaction of Escherichia coli primase with a phage G4ori(c)-E. coli SSB complex

Interaction of Escherichia coli primase with a phage G4ori(c)-E. coli SSB complex

Template-directed pausing in in vitro DNA synthesis by DNA polymerase a from Drosophila melanogaster embryos.

Secondary Structure in Denatured DNA is Responsible for Its Reaction with Antinative DNA Antibodies of Systemic Lupus Erythematosus Sera

Contact Info

Product

Resources

About