Open AccessCorrect Integration of Model Substrates by Ty1 Integrase
Author(s) -
Sharon P. Moore,
David Garfinkel
Publication year - 2000
Publication title -
journal of virology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.617
H-Index - 292
eISSN - 1070-6321
pISSN - 0022-538X
DOI - 10.1128/jvi.74.24.11522-11530.2000
Subject(s) - integrase , biology , nuclease , recombinant dna , plasmid , saccharomyces cerevisiae , transposable element , dna , mobile genetic elements , retrovirus , genetics , transposition (logic) , microbiology and biotechnology , origin of replication , virology , genome , virus , gene , linguistics , philosophy
The retrovirus-like mobile genetic element ofSaccharomyces cerevisiae , Ty1, transposes to new genomic locations via the element-encoded integrase (IN). Here we report that purified recombinant IN catalyzed correct integration of a linear DNA into a supercoiled target plasmid. Ty1 virus-like particles (VLPs) integrated donor DNA more efficiently than IN. VLP and IN-mediated insertions occurred at random sites in the target. Mg2+ was preferred over Mn2+ for correct integration, and neither cation enhanced nonspecific nuclease activity of IN. Products consistent with correct integration events were also obtained by Southern analysis. Recombinant IN and VLPs utilized many, but not all, linear donor fragments containing non-Ty1 ends, including a U3 mutation which has been shown to be defective for transposition in vivo. Together, our results suggest that IN is sufficient for Ty1 integration in vitro and IN interacts with exogenous donors less stringently than with endogenous elements.