Abstract
The coordinated synthesis of both leading and lagging DNA strands is thought to involve a dimeric DNA polymerase and a looping of the lagging strand so that both strands can be synthesized in the same direction. We have constructed a minicircle with a replication fork that permits an assessment of the stoichiometry of the proteins and a measurement of the synthesis of each strand. The replisome consisting of bacteriophage T7 DNA polymerase, helicase, primase, and single-stranded DNA-binding protein mediates coordinated replication. The criteria for coordination are fulfilled: (1) a replication loop is formed, (2) leading and lagging strand synthesis are coupled, (3) the lagging strand polymerase recycles from one Okazaki fragment to another, and (4) the length of Okazaki fragments is regulated. T7 single-stranded DNA-binding protein is essential for coordination.
Publication types
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, P.H.S.
MeSH terms
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Bacteriophage T7 / chemistry
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Bacteriophage T7 / enzymology
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Bacteriophage T7 / genetics
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Base Sequence
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Binding Sites / genetics
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DNA / biosynthesis*
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DNA / chemistry
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DNA / metabolism
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DNA / ultrastructure
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DNA Helicases / metabolism
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DNA Polymerase III / metabolism
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DNA Primase / metabolism
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DNA Replication*
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DNA, Circular / chemistry
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DNA, Circular / metabolism*
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DNA, Circular / ultrastructure
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DNA, Single-Stranded / chemistry
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DNA, Single-Stranded / metabolism
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DNA, Viral / chemistry
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DNA, Viral / metabolism
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DNA-Binding Proteins / genetics
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DNA-Binding Proteins / metabolism
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Molecular Sequence Data
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Nucleic Acid Conformation
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Templates, Genetic
Substances
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DNA, Circular
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DNA, Single-Stranded
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DNA, Viral
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DNA-Binding Proteins
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Okazaki fragments
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DNA
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DNA Primase
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DNA Polymerase III
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DNA Helicases