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Analysis of the N‐terminal DNA binding domain of the IS 30 transposase
Author(s) -
Nagy Zita,
Szabó Mónika,
Chandler Michael,
Olasz Ferenc
Publication year - 2004
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.1365-2958.2004.04279.x
Subject(s) - transposase , insertion sequence , biology , inverted repeat , genetics , mutant , transposable element , sequence alignment , sinorhizobium meliloti , dna , direct repeat , conserved sequence , peptide sequence , gene , genome , base sequence
Summary IS 30 is the founding member of a large family of widely spread bacterial insertion sequences with closely related transposases. The N‐terminal end of the IS 30 transposase had been shown to retain sequence‐specific DNA binding activity and to protect the IS 30 terminal inverted repeats. Structural predictions revealed the presence of a helix–helix–turn–helix motif (H–HTH2) which, in the case of IS 30 , is preceded by an additional helix–turn–helix motif (HTH1). Analysis of deletion and point mutants in this region revealed that both motifs are important for IS 30 transposition. IS 30 exhibits two types of insertion specificity preferring either a 24 bp palindromic hot‐spot (GOHS) or sequences resembling its ends [left and right terminal inverted repeat (IRL and IRR)]. Results are presented suggesting that the HTH1 region is required for GOHS targeting and interferes with the inverted repeat (IR) targeting. On the other hand, H–HTH2 appears to be required for both. The binding activities of the mutant proteins to the terminal IS 30 IRs as measured by gel retardation correlated well with these results. Furthermore, close inspection of the H–HTH2 region revealed significant amino acid identity with a similar predicted secondary structure carried by the transcriptional regulator FixJ of Sinorhizobium meliloti and involved in FixJ binding to its target sequence. This suggests that FixJ and IS 30 transposase share similar sequence‐specific DNA binding mechanisms.