Determinants of Bacteriophage 933W Repressor DNA Binding Specificity

Abstract

We reported previously that 933W repressor apparently does not cooperatively bind to adjacent sites on DNA and that the relative affinities of 933W repressor for its operators differ significantly from that of any other lambdoid bacteriophage. These findings indicate that the operational details of the lysis-lysogeny switch of bacteriophage 933W are unique among lambdoid bacteriophages. Since the functioning of the lysis-lysogeny switch in 933W bacteriophage uniquely and solely depends on the order of preference of 933W repressor for its operators, we examined the details of how 933W repressor recognizes its DNA sites. To identify the specificity determinants, we first created a molecular model of the 933W repressor-DNA complex and tested the predicted protein-DNA interactions. These results of these studies provide a picture of how 933W repressor recognizes its DNA sites. We also show that, opposite of what is normally observed for lambdoid phages, 933W operator sequences have evolved in such a way that the presence of the most commonly found base sequences at particular operator positions serves to decrease, rather than increase, the affinity of the protein for the site. This finding cautions against assuming that a consensus sequence derived from sequence analysis defines the optimal, highest affinity DNA binding site for a protein.

Mechanisms that Determine the Differential Stability of Stx+ and Stx− Lysogens

“…We envision three mechanisms could cause this decreased occupancy: (1) a decreased DNA binding affinity of Stx + phage repressors for their respective operators, (2) an increased ability of Stx + phage repressors to undergo self-cleavage, and/or (3) a decreased total amount of Stx + phage repressors present in the lysogens [ 27 ]. Our work [ 27 , 32 ] indicates that the affinities of the Stx + phage repressors for their DNA sites are similar or higher than the affinities of other lambdoid phage repressors for their cognate operators [ 37 , 38 , 39 , 40 , 41 ]. While we have not directly compared the sensitivity of 933W, BAA2326 , 434, and λ repressors to RecA-mediated autocleavage, our results ( Figure 5 ) indicate that the increased sensitivity of the Stx + lysogens to spontaneous induction is due, at least in part, to a lower amount of repressor in the 933W lysogen than found in λ lysogens.…”

Genes from the exo–xis region of λ and Shiga toxin-converting bacteriophages influence lysogenization and prophage induction

“…Therefore, understanding of mechanisms of regulation of this step in phage development is important for both basic knowledge and development of potential anti-STEC therapies. It is worth mentioning that although principles of the prophage induction have been described for cells lysogenized with bacteriophage λ and cultured under laboratory conditions (for reviews see Ptashne 2004 ; Węgrzyn and Węgrzyn 2005 ) and despite the fact that recent reports provided important information about regulation of lysogenization by Stx phages and induction of corresponding prophages (Aertsen et al 2005 ; Bullwinkle and Koudelka 2011 ; Bullwinkle et al 2012 ; Fogg et al 2007 , 2010 , 2011 , 2012 ; Imamovic and Muniesa 2012 ; Łoś et al 2009 , 2010 ; Murphy et al 2008 ; Nejman et al 2009 , 2011 ; Nejman-Faleńczyk et al 2012 ; Riley et al 2012 ; Smith et al 2012 ), our knowledge on modulation of these processes by various factors and conditions is still far from completeness.…”

Genomic Characterization of Two Shiga Toxin–Converting Bacteriophages Induced From Environmental Shiga Toxin–Producing Escherichia coli

“…Stx prophages can be induced from STEC strains by different external stresses and subsequently released in the environment as Stx-converting phages, which are capable of infecting other susceptible strains and pose a risk of virulence gene transfer. Bacteriophage 933W, a well-studied Stx-converting phage, can self-induce from E. coli O157:H7 EDL933 strain and subsequently infect other susceptible E. coli strains to form a new lysogen ( Bullwinkle et al, 2012 ). Stx-converting phages are also highly associated with the evolution of pathogens and result in the increased pathogenicity of these pathogens.…”