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The phage lambda major tail protein structure reveals a common evolution for long-tailed phages and the type VI bacterial secretion system - PubMed

  • ️Thu Jan 01 2009

The phage lambda major tail protein structure reveals a common evolution for long-tailed phages and the type VI bacterial secretion system

Lisa G Pell et al. Proc Natl Acad Sci U S A. 2009.

Abstract

Most bacteriophages possess long tails, which serve as the conduit for genome delivery. We report the solution structure of the N-terminal domain of gpV, the protein comprising the major portion of the noncontractile phage lambda tail tube. This structure is very similar to a previously solved tail tube protein from a contractile-tailed phage, providing the first direct evidence of an evolutionary connection between these 2 distinct types of phage tails. A remarkable structural similarity is also seen to Hcp1, a component of the bacterial type VI secretion system. The hexameric structure of Hcp1 and its ability to form long tubes are strikingly reminiscent of gpV when it is polymerized into a tail tube. These data coupled with other similarities between phage and type VI secretion proteins support an evolutionary relationship between these systems. Using Hcp1 as a model, we propose a polymerization mechanism for gpV involving several disorder-to-order transitions.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.
Fig. 1.

Solution structure of gpVN. (A) A ribbon representation of the lowest energy structure of gpVN. (B) The 16 hydrophobic core side-chains are represented as yellow bonds. Strands are colored in dark blue, the helix in red, and loops in gray. A and B were generated by using PyMOL (

http://pymol.sourceforge.net/

).

Fig. 2.
Fig. 2.

Alignment of diverse homologues of gpV. The gpVN homologues shown here are representative of many sequences collected through many iterations of PSI-BLAST (for details, see Methods). These sequences were chosen to maximize diversity, and are derived mostly from prophages in various bacteria (sequences are designated by National Center for Biotechnology Information GI nos.). Sequences taken from characterized phages are indicated. The average pairwise identity of these sequences to gpVN is 17%, with no sequence being >23% identical. The average pairwise identity among all of the sequences in the alignment is 18%. No 2 sequences in the alignment are >37% identical. Nonconserved sequences at the N termini of these proteins were truncated (gpV starts at Met 8). In the last 2 sequences, a large loop after β-strand 4 is truncated. The 14 highly conserved hydrophobic core positions are indicated in yellow. Conserved nonpolar positions in the structured regions of gpVN that are >20% exposed in the gpVN monomeric structure are indicated in red. Conserved residues within the middle unstructured loop are indicated in orange, and additional residues that are conserved and found at hexamer interface positions are indicated in cyan. Experimentally derived secondary structure elements are indicated on top of the alignment in blue with the computationally predicted secondary structure shown in yellow (15).

Fig. 3.
Fig. 3.

Role of the central disordered loop in λ tail assembly. (A) The ability of the D61A/D62A gpV mutant expressed from a plasmid to complement a λ Vam lysate in vivo was assessed by observing plaque formation. The “empty vector” experiment constitutes the negative control. As indicated, 3 different dilutions of the Vam lysate were spotted onto the plates. (B) The effect of plasmid-expressed WT gpV and the D61A/D62A mutant on the growth of WT λ phage was assayed. A WT phage lysate was spotted at the indicated dilutions on cells carrying these plasmids. (C) Representative electron micrographs of phage particles produced when a WT λ prophage was induced in the presence of empty vector, overexpressed wild-type gpV, or overexpressed D61A/D62A gpV. (Scale bar, 100 nm.)

Fig. 4.
Fig. 4.

Structural similarity between gpVN and a TTP from a contractile-tailed phage. (A) Gene order conservation among tail proteins from λ (Siphoviridae), HK97 (Siphoviridae), P2 (Myoviridae), Mu (Myoviridae), and PBSX (Myoviridae). In each case, the tail chaperones (gpG and frameshift product, gpGT, in λ), colored purple, are located between the gene encoding the MTP or TTP, colored blue, and the gene encoding the TMP, colored orange. In Myoviridae, the gene encoding the TSP, colored red, is located upstream of the TTP. It should be noted that PBSX was previously reported to possess the conserved frameshift (9), but this was an error. (B) Tertiary and (C) secondary structure comparison between gpVN and XkdM, the TTP from PBSX. Each secondary structure element in XkdM is color-coded based on its tertiary alignment with gpVN (β1, salmon; β2, orange; β3, yellow; β4, blue; α1, green; β5, sky blue; β6, purple; and β7, deep pink). Loops and secondary structure elements that do not align are colored in gray. In C, regions that are disordered in gpVN are denoted with a red star.

Fig. 5.
Fig. 5.

Structural similarities between gpVN and Hcp1. (A) Tertiary and (B) secondary structure comparison between gpVN and Hcp1. Each secondary structure element in Hcp1 is colored-coded based on its alignment with gpVN (β1, salmon; β2, orange; β3, yellow; β4, blue; α1, green; β5, sky blue; β6, purple; and β7, deep pink). Loops and secondary structure differences are colored in gray, with the exception of the β2-β3 loop that is colored red. In B, disordered regions in gpVN are denoted with a red star. (C) Superposition of gpV (blue) onto monomer A of the Hcp1 hexamer (yellow). Crystallographic symmetry was applied to build the stacked Hcp1 hexamers. The β2-β3 loop of Hcp1 is represented in red spheres, which are clearly located in the hexamer-hexamer interface. (D) Structural superpositions suggest that the disordered C-terminal residues of gpVN could form a strand. For ease of visualization, in the superposition of Hcp1 and gpVN, residues 32–56 and 138–162 from Hcp1 and 49–82 from gpVN have been removed, and in the superposition of XkdM with gpVN, residues 33–59 from XkdM and 49–82 from gpVN have been removed. The folding of the gpVN C terminus (deep pink) into the Hcp1 β8′ (deep pink) or the XkdM β7 position (deep pink) are likely conformational changes. (E) A side view of 3 monomers from the Hcp1 hexamer where monomer A has been replaced with gpVN (blue). Highly conserved hydrophobic residues are colored red, whereas relatively conserved residues are colored in cyan. Conserved residues in the β2-β3 loop are orange. Residues contributing to the hexameric interface are circled in black (Asp-44, Trp-86, Gln-93, Trp-123, Phe-112, Phe-120, Val-136, and Ile-137).

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