Functional Evolution of the Cro Protein Family of Transcription Factors
AdvisorCordes, Matthew H
Committee ChairCordes, Matthew H
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PublisherThe University of Arizona.
RightsCopyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author.
AbstractMembers of multi-specific DNA-binding protein families have evolved to specifically recognize diverse DNA site sequences. This dissertation presents evidence that the Cro protein family of helix-turn-helix transcription factors from lambdoid bacteriophages may share a conserved, limited "code" that partially governs evolution of their binding specificity. A bioinformatic study revealed six conserved sequence correlations between residues at three positions in Cro recognition helices and three base-pairs in putative cognate DNA consensus half-sites (Chapter 2). Three of these pairings correspond to sequence-specific contacts observed at the binding interface of lambda Cro and consensus operator DNA in a previously available co-crystal structure (Albright and Matthews, 1998a). In vitro mutagenesis and functional characterization was used to validate the proposed "code" (Chapter 3). Two out of three "coding" combinations acted as specificity switches in lambda Cro, though variant proteins displayed reduced binding specificity for their predicted target DNA sites. Two crystal structures of a lambda Cro variant are presented in Chapter 4, which provide insight into lambda Cro dimer flexibility. Additionally, a co-crystal structure of N15 Cro bound to consensus site DNA was determined which contains two coding residue pairs at the binding interface (Chapter 5), and a crystal structure of Xfasa1 Cro that enables future investigations into Cro functional evolution (Chapter 6). Although there are several caveats, the data are consistent with a model in which Cro proteins may indeed have evolved new binding specificities in part through simple mutations at their binding interfaces that follow a simple set of evolutionarily conserved "coding" rules. The structural and functional diversity of Cro proteins provides an exciting venue for future research into their evolution.
Degree ProgramBiochemistry & Molecular Biophysics