Differential role of the Mu B protein in phage Mu integration vs. replication: mechanistic insights into two transposition pathways.
The Mu B protein is an ATP-dependent DNA-binding protein and an allosteric activator of the Mu transposase. As a result of these activities, Mu B is instrumental in efficient transposition and target-site choice. We analysed in vivo the role of Mu B in the two different recombination reactions performed by phage Mu: non-replicative transposition, the pathway used during integration, and replicative transposition, the pathway used during lytic growth. Utilizing a sensitive PCR-based assay for Mu transposition, we found that Mu B is not required for integration, but enhances the rate and extent of the process. Furthermore, three different mutant versions of Mu B, Mu BC99Y, Mu BK106A, and Mu B1-294, stimulate integration to a similar level as the wild-type protein. In contrast, these mutant proteins fail to support Mu growth. This deficiency is attributable to a defect in formation of an essential intermediate for replicative transposition. Biochemical analysis of the Mu B mutant proteins reveals common features: the mutants retain the ability to stimulate transposase, but are defective in DNA binding and target DNA delivery. These data indicate that activation of transposase by Mu B is sufficient for robust non-replicative transposition. Efficient replicative transposition, however, demands that the Mu B protein not only activate transposase, but also bind and deliver the target DNA.
Roldan
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Department of Biology and the Howard Hughes Medical Institute, Massachusetts Institute of Technology, 77 Massachusetts Avenue, 68-523 Cambridge, MA 02139, USA.
Baker
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TA
D010583:Bacteriophage mu / Q000502:physiology*; D001483:Base Sequence; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000502:physiology*; D014764:Viral Proteins; D016662:Virus Integration / Q000502:physiology*; D014779:Virus Replication / Q000502:physiology*
D017931:DNA Primers; D004268:DNA-Binding Proteins; C050938:Mu B protein, bacteriophage; D014764:Viral Proteins