Mutations of the phage lambda attachment site alter the directionality of resolution of Holliday structures.

AUTOR(ES)

de Massy, B

RESUMO

Integrative recombination of bacteriophage lambda occurs by two sequential, reciprocal strand exchanges at specific positions within the attachment sites. Both exchanges are promoted by the lambda Int protein; the first forms a Holliday structure, and the second resolves it to recombinant products. Recombination requires sequence homology within the 7 bp 'overlap' region that separates the two points of strand exchange. To see if homology promotes the second strand exchange, we constructed attachment site Holliday structures by annealing DNA strands and then assayed Int-promoted resolution. Holliday structures corresponding to strand exchange between sites with homologous overlap regions were efficiently resolved to give mixtures of recombinants and parents. Holliday structures corresponding to exchanges between heterologous sites fell into two classes. Members of the first class, in which heterology limited but did not completely prevent migration of the branchpoint within the overlap region, were resolved efficiently and preferentially to parental molecules. We propose that resolution to recombinants occurs only if homology allows branch migration from the first to the second exchange site. Members of the second class, in which heterology constrained the branchpoint within an Int binding site, were resolved poorly. We suggest that Holliday structures that have a branchpoint within an Int binding site are poor substrates for Int.

Documentos Relacionados

• The extent of DNA sequence required for a functional bacterial attachment site of phage lambda.
• Dissecting the resolution reaction of lambda integrase using suicide Holliday junction substrates.
• Isolation of point mutations in bacteriophage Mu attachment regions cloned in a lambda::mini-Mu phage.
• Resolution of synthetic Holliday structures by an extract of human cells.
• Interaction of the lambda site-specific recombination protein Xis with attachment site DNA.