Analysis of the lambdoid prophage element e14 in the E. coli K-12 genome

Abstract

Background: Many sequenced bacterial genomes harbor phage-like elements or cryptic prophages. These elements have been implicated in pathogenesis, serotype conversion and phage immunity. The e14 element is a defective lambdoid prophage element present at 25 min in the E. coli K-12 genome. This prophage encodes important functional genes such as lit (T4 exclusion), mcrA (modified cytosine restriction activity) and pin (recombinase).

Results: Bioinformatic analysis of the e14 prophage sequence shows the modular nature of the e14 element which shares a large part of its sequence with the Shigella flexneri phage SfV. Based on this similarity, the regulatory region including the repressor and Cro proteins and their binding sites were identified. The protein product of b1149 was found to be a fusion of a replication protein and a terminase. The genes b1143, b1151 and b1152 were identified as putative pseudogenes. A number of duplications of the stfE tail fibre gene of the e14 are seen in plasmid p15B. A protein based comparative approach using the COG database as a starting point helped detect lambdoid prophage like elements in a representative set of completely sequenced genomes.

Conclusions: The e14 element was characterized for the function of its encoded genes, the regulatory regions, replication origin and homology with other phage and bacterial sequences. Comparative analysis at nucleotide and protein levels suggest that a number of important phage related functions are missing in the e14 genome including parts of the early left operon, early right operon and late operon. The loss of these genes is the result of at least three major deletions that have occurred on e14 since its integration. A comparative protein level approach using the COG database can be effectively used to detect defective lambdoid prophage like elements in bacterial genomes.

Figure 1

Overview of the e14 genome. The genetic functions of a generic lambdoid bacteriophage genome (brown rectangle) are shown above displayed with a transcriptional map (black arrows). In the middle, the section of the E. coli K-12 genome that contains e14 (gray rectangle) is shown with ORFs denoted by rectangular arrows oriented in the direction of transcription (green – host genes; red – e14 genes that are likely nonfunctional; black – e14 genes that are known to be functional; blue – e14 genes whose functionality cannot be assessed at present; parentheses indicate the boundaries of the P-invertable element). Small black arrows above the e14 map denote putative promoters, vertical lines denote putative terminators and small black squares putative operators. The yellow regions between the lambdoid and e14 maps indicate regions where e14 has homology to at least one known member of the lambdoid phage family (see text for details). Below, colored rectangles mark regions of highest homology between e14 and various known phages and prophages with regions of greater similarity closer to the e14 map (these are not meant to show all known homologies, only the closest ones); CPS-53 is a defective prophage in E. coli K-12, CP-933H is prophage in E. coli EDL933 and CP073-5, Sti4b, and Sti8 are provisional names for prophages in E. coli CFT073 and S. typhi CT18 (Supplementary Material of Ref. [39]).

Figure 2

The regulatory region of the e14 element. The possible ORFs for the cro (b1146) and cI repressors (b1145) are indicated in blue and orientation indicated by the arrow. The inverted repeats as detected by Allison et al. [31] for SfV are boxed. The palindromic regions are underlined. The ribosome binding sites (RBS) and putative -10 and -35 for the early right operon are indicated by different color letters within the box.

Figure 3

Cumulative GC-plot of e14. Cumulative GC-plot of e14 using a window size of 500 showing the regions of minima, which were analyzed for possible origins of replication. The y-axis represents ∑ (G-C)/(G+C) multiplied by 1000. The x-axis gives the base positions in e14.