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Allosteric regulation of the primase (DnaG) activity by the clamp‐loader (τ) in vitro
Author(s) -
Chintakayala Kiran,
Machón Cristina,
Haroniti Anna,
Larson Marilyn A.,
Hinrichs Steven H.,
Griep Mark A.,
Soultanas Panos
Publication year - 2009
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1111/j.1365-2958.2009.06668.x
Subject(s) - dnab helicase , dnag , primase , replisome , biology , helicase , dna replication , dna polymerase , biochemistry , dna , circular bacterial chromosome , rna , reverse transcriptase , gene
Summary During DNA replication the helicase (DnaB) recruits the primase (DnaG) in the replisome to initiate the polymerization of new DNA strands. DnaB is attached to the τ subunit of the clamp‐loader that loads the β clamp and interconnects the core polymerases on the leading and lagging strands. The τ–DnaB−DnaG ternary complex is at the heart of the replisome and its function is likely to be modulated by a complex network of allosteric interactions. Using a stable ternary complex comprising the primase and helicase from Geobacillus stearothermophilus and the τ subunit of the clamp‐loader from Bacillus subtilis we show that changes in the DnaB–τ interaction can stimulate allosterically primer synthesis by DnaG in vitro . The A550V τ mutant stimulates the primase activity more efficiently than the native protein. Truncation of the last 18 C‐terminal residues of τ elicits a DnaG‐stimulatory effect in vitro that appears to be suppressed in the native τ protein. Thus changes in the τ–DnaB interaction allosterically affect primer synthesis. Although these C‐terminal residues of τ are not involved directly in the interaction with DnaB, they may act as a functional gateway for regulation of primer synthesis by τ‐interacting components of the replisome through the τ–DnaB−DnaG pathway.