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Roles of the helicase and primase domain of the gene 4 protein of bacteriophage T7 in accessing the primase recognition site
Author(s) -
Kusakabe Takahiro,
Baradaran Khandan,
Lee Joonsoo,
Richardson Charles C.
Publication year - 1998
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.1093/emboj/17.5.1542
Subject(s) - primase , biology , helicase , genetics , bacteriophage , gene , microbiology and biotechnology , polymerase chain reaction , reverse transcriptase , rna , escherichia coli
The 63 kDa gene 4 protein of bacteriophage T7 provides both helicase and primase activities. The C‐terminal helicase domain of the gene 4 protein is responsible for DNA‐dependent NTP hydrolysis and for hexamer formation, whereas the N‐terminal primase domain contains the zinc motif that is, in part, responsible for template‐directed oligoribonucleotide synthesis. In the presence of β,γ‐methylene dTTP, the protein forms a hexamer that surrounds and binds tightly to single‐stranded DNA and consequently is unable to translocate to primase recognition sites, 5′‐GTC‐3′, or to dissociate from the molecule to which it is bound. Nonetheless, in the presence of β,γ‐methylene dTTP, it catalyzes the synthesis of pppAC dimers at primase sites on M13 DNA. When bound to single‐stranded DNA in the presence of β,γ‐methylene dTTP, the primase can function at recognition sites on the same molecule to which it is bound provided that a sufficient distance exists between the recognition site and the site to which it is bound. Furthermore, the primase bound to one DNA strand can function at a primase site located on a second DNA strand. The results indicate that the primase domain resides on the outside of the hexameric ring, a location that enables it to access sites distal to its site of binding.