Brujita Integrase: A Simple, Arm-Less, Directionless, and Promiscuous Tyrosine Integrase System.
Mycobacteriophage Brujita is an unusual temperate phage in which establishment of superinfection immunity is dependent on chromosomal integration. Integration is mediated by a non-canonical tyrosine integrase (Int) lacking an N-terminal domain typically associated with binding to arm-type sites within the phage attachment site (attP). This raises the question as to how these Ints bind their DNA substrates, if they form higher-order protein DNA complexes, and how site selection and recombinational directionality are determined. Here we show that Brujita Int is a simple recombinase, whose properties more closely resemble those of FLP and Cre than it does the canonical phage Ints. Brujita Int uses relatively small DNA substrates, fails to discriminate between attP and attB, cleaves attachment site DNA to form a 6-base overlap region, and lacks directional control. Brujita Int also has an unusual pattern of binding to its DNA substrates. It binds to two half sites (B and B') at attB, although binding to the B half site is strongly dependent on occupancy of B'. In contrast, binding to the P half site is not observed, even when Int is bound at P'. However, an additional Int binding site (P1) is displaced to the left of the crossover site at attP, is required for recombination and is predicted to facilitate binding of Int to the P half site during synapsis. These simple phage Int systems may reflect ancestral states of phage evolution with the complexities of higher-order complex formation and directional control representing subsequent adaptations.
Department of Biological Sciences, University of Pittsburgh, Pittsburgh, PA 15260, USA.
Bryce L
BL
Lunt
Department of Biological Sciences, University of Pittsburgh, Pittsburgh, PA 15260, USA. Electronic address: gfh@pitt.edu.
Graham F
GF
Hatfull
D001287:Attachment Sites, Microbiological / Q000235:genetics; D001435:Bacteriophages / Q000235:genetics; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D004247:DNA / Q000235:genetics*; D019426:Integrases / Q000235:genetics*; D045522:Recombinases / Q000235:genetics; D011995:Recombination, Genetic / Q000235:genetics; D014443:Tyrosine / Q000235:genetics*
D045522:Recombinases; D014443:Tyrosine; D004247:DNA; D019426:Integrases
phage integrase; tyrosine recombinase