Replication of single-stranded DNA templates by primase-polymerase complexes of the yeast,<i>Saccharomyces cerevisiae</i>

Biswas, Esther E.; Biswas, Subhasis B.

doi:10.1093/nar/16.14.6411

Cited by 11 publications

(13 citation statements)

References 23 publications

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“…The majority of studies have utilized partiallypurified enzymes and factors to reconstitute DNA synthetic activity [Fairman and Stillman, 1988;reviewed in Kelly, 1988;Stillman, 1989;Hurwitz et al, 1990;Tsurimoto et al, 1990]. Although some data exist on the functional association of several enzymes and factors [Mathews and Slabaugh, 1986;Vishwanatha et al, 1986;Tubo and Berezney, 1987;Biswas and Biswas, 1988;Syvä oja and Linn, 1989;Reddy and Fager, 1993;Maga and Hü bscher, 1996], the precise molecular interactions that occur between the proteins required to support DNA replication remain largely undefined. The ability to reconstitute DNA replication activity utilizing individual components does not exclude the existence of large multiprotein complexes containing many or all of the proteins that function in vivo to replicate DNA.…”

Section: Discussionmentioning

confidence: 99%

“…These proteins include DNA polymerases a and d, DNA topoisomerases I and II, DNA primase, DNA ligase I, proliferating cell nuclear antigen (PCNA), replication factor C (RFC), replication protein A (RPA), and the nucleases RNase H1 and FEN1/RTH1 [reviewed in Stillman, 1994;Hickey and Malkas, 1997;Malkas, 1998]. Increasing evidence has supported the concept that many of the enzymes and factors involved in the replication of mammalian DNA function together as an organized multiprotein complex [Baril et al, 1983;Jackson and Cook, 1986;Mathews and Slabaugh, 1986;Vishwanatha et al, 1986;Tubo and Berezney, 1987;Biswas and Biswas, 1988;Syvä oja and Linn, 1989;Reddy and Fager, 1993;Maga and Hü bscher, 1996;Hickey and Malkas, 1997;Malkas, 1998]. We have previously reported that a DNA replication-competent multiprotein form of DNA polymerase can be isolated from a variety of mammalian cell lines and tissues [Malkas et al, 1990;Wu et al, 1994;Applegren et al, 1995;Coll et al, 1996Coll et al, , 1997Tom et al, 1996;Lin et al, 1997].…”

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confidence: 99%

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Human cell DNA replication is mediated by a discrete multiprotein complex

Jiang

Hickey

Abdel-Aziz

et al. 2002

J of Cellular Biochemistry

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A discrete high molecular weight multiprotein complex containing DNA polymerase alpha has been identified by a native Western blotting technique. An enrichment of this complex was seen at each step in its purification. Further purification of this complex by ion-exchange chromatography indicates that the peak of DNA polymerase alpha activity co-purifies with the peak of in vitro SV40 DNA replication activity eluting from the column. The complex has a sedimentation coefficient of 18S in sucrose density gradients. We have designated this complex as the DNA synthesome. We further purified the DNA synthesome by electroeluting this complex from a native polyacrylamide gel. The eluted complex retains in vitro DNA synthetic activity, and by Western blot analysis, contains DNA polymerase delta, proliferating cell nuclear antigen, and replication protein A. Enzymatic analysis of the electroeluted DNA synthesome indicates that the synthesome contains topoisomerase I and II activities, and SDS-PAGE analysis of the electroeluted DNA synthesome revealed the presence of at least 25 major polypeptides with molecular weights ranging from 20 to 240 kDa. Taken together, our evidence suggests that the DNA synthesome may represent the minimal DNA replication unit of the human cell.

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Section: Discussionmentioning

confidence: 99%

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confidence: 99%

Human cell DNA replication is mediated by a discrete multiprotein complex

Jiang

Hickey

Abdel-Aziz

et al. 2002

J of Cellular Biochemistry

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“…M13-ARS, containing an 837-base-pair (bp) HindIII-EcoRI fragment of ARS1, was subcloned in M13SK phagemid (Stratagene, La Jolla, Calif.) as previously described (3). Construction of TF1-ARS and TF5-ARS plasmids was as previously described (10,11).…”

Section: Cm)mentioning

confidence: 99%

ARS binding factor I of the yeast Saccharomyces cerevisiae binds to sequences in telomeric and nontelomeric autonomously replicating sequences.

Biswas¹,

Biswas²

1990

Mol. Cell. Biol.

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We have analyzed various autonomously replicating sequences (ARSs) in yeast nuclear extract with ARS-specific synthetic oligonucleotides. The El oligonucleotide sequence, which is derived from HMRE-ARS, and the F1 oligonucleotide sequence, which is derived from telomeric ARS120, appeared to bind to the same cellular factor with high specificity. In addition, each of these oligonucleotides was a competitive inhibitor of the binding of the other. Binding of the ARS binding factor (ABF) to either of these oligonucleotides was inhibited strongly by plasmids containing ARSI and telomeric TF1-ARS. DNase I footprinting analyses with yeast nuclear extract showed that El and F1 oligonucleotides eliminated protection of the binding site of ARS binding factor I (ABFI) in domain B of ARSi. Sequence analyses of various telomeric (ARS120 and TFl-ARS) and nontelomeric ARSs (ARSi and HMRE-ARS) showed the presence of consensus ABFI binding sites in the protein binding domains of all of these ARSs. Consequently, the ABFI and ABFI-like factors bind to these domain B-like sequences in a wide spectrum of ARSs, both telomeric and nontelomeric.Sequence-specific DNA binding has been shown to be involved in the control of DNA replication and transcription in both procaryotic and eucaryotic systems (1-9, 15-17, 27-31, 34-38). The sequence-specific DNA binding of cellular factors presumably controls the initiation of DNA replication in these origin sites in a fashion similar to that observed with Escherichia coli, simian virus 40, and bacteriophage lambda DNA replication (1,2,15,16,22,27,33). In E. coli, the DnaA protein binds to specific sequences at the origin of DNA replication (oriC) and allows initiation of chromosomal DNA replication (22). The genome of the yeast

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“…We identified earlier a high molecular weight form of primase-polymerase complex from Saccharomyces cerevisiae which was capable of rapid and processive DNA synthesis on an unprimed single-stranded DNA template (Biswas & Biswas, 1988). In this paper, we describe the purification of a multimeric form of DNA polymerase a to homogeneity.…”

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“…These four subunits may likely be a part of a larger multiprotein complex that represents the replicating form of polymerase a. In fact, studies in several laboratories have indicated that DNA polymerase a could be isolated as a large complex accompanied by a number of enzymatic activities such as polymerase, primase, and exonuclease (Hubscher et al, 1982;Baril et al, 1983; Takada et al, 1986;; Biswas & Biswas, 1988). The multimeric form of t This work was supported by a grant from the U.S. Public Health Service.…”

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Purification and characterization of a yeast DNA polymerase .alpha. complex with associated primase, 5'.fwdarw.3' exonuclease, and DNA-dependent ATPase activities

Biswas

Chen²,

Gray³

et al. 1993

Biochemistry

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We have purified a multimeric form of yeast DNA polymerase alpha with DNA polymerase, primase, 5'-->3' exonuclease, and single-stranded (ss) DNA-dependent ATPase activities to near-homogeneity. The molecular mass of complex was 650 kDa with subunits ranging in sizes from 30 to 180 kDa. The alpha-subunit of the complex could be detected by DNA polymerase alpha antibody. No cross-reactivity of polypeptides within the complex was observed with antibodies directed against polymerase delta or epsilon. The multimeric polymerase alpha could be selectively inhibited by p-n-butylphenyl-dGTP (I50 of approximately 0.2 microM), p-n-butylanilino-dATP (I50 of 1.3 microM), and aphidicolin (I50 of 2.5 micrograms/mL). The complex synthesized RNA primers on various ssDNA templates and rapidly elongated these primers into nascent DNA fragments in the presence of required deoxynucleotides. It has a strong 5'-->3' exonuclease activity. In addition, the complex hydrolyzed both ATP and dATP in a ssDNA-dependent manner. Thus, the multiprotein complex of DNA polymerase alpha had multiple activities (primase, polymerase, and ATPase) which could act concertedly to synthesize primers and elongate the primers to nascent DNA fragments in the lagging strand of the fork.

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Replication of single-stranded DNA templates by primase-polymerase complexes of the yeast,Saccharomyces cerevisiae

Cited by 11 publications

References 23 publications

Human cell DNA replication is mediated by a discrete multiprotein complex

Human cell DNA replication is mediated by a discrete multiprotein complex

ARS binding factor I of the yeast Saccharomyces cerevisiae binds to sequences in telomeric and nontelomeric autonomously replicating sequences.

Purification and characterization of a yeast DNA polymerase .alpha. complex with associated primase, 5'.fwdarw.3' exonuclease, and DNA-dependent ATPase activities

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