A quantitative binding model for the Apl protein, the dual purpose recombination-directionality factor and lysis-lysogeny regulator of bacteriophage 186

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dc.contributor.authorCutts, E.E.
dc.contributor.authorEgan, J.
dc.contributor.authorDodd, I.B.
dc.contributor.authorShearwin, K.E.
dc.date.issued2020
dc.description.abstractThe Apl protein of bacteriophage 186 functions both as an excisionase and as a transcriptional regulator; binding to the phage attachment site (att), and also between the major early phage promoters (pR-pL). Like other recombination directionality factors (RDFs), Apl binding sites are direct repeats spaced one DNA helix turn apart. Here, we use in vitro binding studies with purified Apl and pR-pL DNA to show that Apl binds to multiple sites with high cooperativity, bends the DNA and spreads from specific binding sites into adjacent non-specific DNA; features that are shared with other RDFs. By analysing Apl's repression of pR and pL, and the effect of operator mutants in vivo with a simple mathematical model, we were able to extract estimates of binding energies for single specific and non-specific sites and for Apl cooperativity, revealing that Apl monomers bind to DNA with low sequence specificity but with strong cooperativity between immediate neighbours. This model fit was then independently validated with in vitro data. The model we employed here is a simple but powerful tool that enabled better understanding of the balance between binding affinity and cooperativity required for RDF function. A modelling approach such as this is broadly applicable to other systems.
dc.description.statementofresponsibilityErin E Cutts, J Barry Egan, Ian B Dodd, Keith E Shearwin
dc.identifier.citationNucleic Acids Research, 2020; 48(16):8914-8926
dc.identifier.doi10.1093/nar/gkaa655
dc.identifier.issn0305-1048
dc.identifier.issn1362-4962
dc.identifier.orcidDodd, I.B. [0000-0003-2969-6841]
dc.identifier.orcidShearwin, K.E. [0000-0002-7736-2742]
dc.identifier.urihttp://hdl.handle.net/2440/128706
dc.language.isoen
dc.publisherOxford University Press
dc.relation.granthttp://purl.org/au-research/grants/arc/DP150103009
dc.relation.granthttp://purl.org/au-research/grants/arc/DP110100824
dc.rights© The Author(s) 2020. Published by Oxford University Press on behalf of Nucleic Acids Research. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.
dc.source.urihttps://doi.org/10.1093/nar/gkaa655
dc.subjectMyoviridae
dc.subjectDNA Nucleotidyltransferases
dc.subjectDNA-Binding Proteins
dc.subjectViral Proteins
dc.subjectDNA, Viral
dc.subjectLysogeny
dc.subjectRecombination, Genetic
dc.subjectBinding Sites
dc.subjectProtein Binding
dc.subjectAttachment Sites, Microbiological
dc.subjectPromoter Regions, Genetic
dc.titleA quantitative binding model for the Apl protein, the dual purpose recombination-directionality factor and lysis-lysogeny regulator of bacteriophage 186
dc.typeJournal article
pubs.publication-statusPublished

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