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Collagen-like proteins in pathogenic E. coli strains.

Ghosh N, McKillop TJ, Jowitt TA, Howard M, Davies H, Holmes DF, Roberts IS, Bella J - PLoS ONE (2012)

Bottom Line: Under the electron microscope, collagen-like proteins from E. coli O157:H7 show a dumbbell shape, with two globular domains joined by a hinged stalk.This morphology is consistent with their likely role as trimeric phage side-tail proteins that participate in the attachment of phage particles to E. coli target cells, either directly or through assembly with other phage tail proteins.Thus, collagen-like proteins in enterohaemorrhagic E. coli genomes may have a direct role in the dissemination of virulence-related genes through infection of harmless strains by induced bacteriophages.

View Article: PubMed Central - PubMed

Affiliation: Manchester Interdisciplinary Biocentre, University of Manchester, Manchester, United Kingdom.

ABSTRACT
The genome sequences of enterohaemorrhagic E. coli O157:H7 strains show multiple open-reading frames with collagen-like sequences that are absent from the common laboratory strain K-12. These putative collagens are included in prophages embedded in O157:H7 genomes. These prophages carry numerous genes related to strain virulence and have been shown to be inducible and capable of disseminating virulence factors by horizontal gene transfer. We have cloned two collagen-like proteins from E. coli O157:H7 into a laboratory strain and analysed the structure and conformation of the recombinant proteins and several of their constituting domains by a variety of spectroscopic, biophysical, and electron microscopy techniques. We show that these molecules exhibit many of the characteristics of vertebrate collagens, including trimer formation and the presence of a collagen triple helical domain. They also contain a C-terminal trimerization domain, and a trimeric α-helical coiled-coil domain with an unusual amino acid sequence almost completely lacking leucine, valine or isoleucine residues. Intriguingly, these molecules show high thermal stability, with the collagen domain being more stable than those of vertebrate fibrillar collagens, which are much longer and post-translationally modified. Under the electron microscope, collagen-like proteins from E. coli O157:H7 show a dumbbell shape, with two globular domains joined by a hinged stalk. This morphology is consistent with their likely role as trimeric phage side-tail proteins that participate in the attachment of phage particles to E. coli target cells, either directly or through assembly with other phage tail proteins. Thus, collagen-like proteins in enterohaemorrhagic E. coli genomes may have a direct role in the dissemination of virulence-related genes through infection of harmless strains by induced bacteriophages.

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Related in: MedlinePlus

Analysis by analytical ultracentrifugation of the average molar mass of a sample of purified rEPclA as a function of increasing concentration of guanidinium chloride (GuHCl).Weight-averaged molar mass was determined using a single ideal species model (see Methods). Mean value masses for the upper and lower plateaux were 138±6 kDa and 43±1 kDa respectively (averages of three measures). The molecular mass of native rEPclA (0 M GuHCl) is consistent with three times that of denatured rEPclA (see text). The transition midpoint concentration is 2.38±0.09 M GuHCl.
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pone-0037872-g003: Analysis by analytical ultracentrifugation of the average molar mass of a sample of purified rEPclA as a function of increasing concentration of guanidinium chloride (GuHCl).Weight-averaged molar mass was determined using a single ideal species model (see Methods). Mean value masses for the upper and lower plateaux were 138±6 kDa and 43±1 kDa respectively (averages of three measures). The molecular mass of native rEPclA (0 M GuHCl) is consistent with three times that of denatured rEPclA (see text). The transition midpoint concentration is 2.38±0.09 M GuHCl.

Mentions: The quaternary structure of rEPclA was investigated by sedimentation equilibrium analytical ultracentrifugation (AUC) at increasing concentrations of guanidinium chloride (GuHCl) (Figure 3). The relative molar mass of rEPclA at 0 M GuHCl was 138±6 kDa, corresponding to the predicted molecular weight of a trimer of rEPclA molecules (3×47 kDa). As the concentration of GuHCl increased, a transition from trimer to monomer was observed and the relative molar mass of rEPclA at 5 M GuHCl was 43 kDa, which is consistent with the predicted molecular weight of the rEPclA monomer. Thus, rEPclA trimers dissociate into monomers as the GuHCl concentration increases; the trimer-to-monomer transition point was estimated at around 2.5 M GuHCl.


Collagen-like proteins in pathogenic E. coli strains.

Ghosh N, McKillop TJ, Jowitt TA, Howard M, Davies H, Holmes DF, Roberts IS, Bella J - PLoS ONE (2012)

Analysis by analytical ultracentrifugation of the average molar mass of a sample of purified rEPclA as a function of increasing concentration of guanidinium chloride (GuHCl).Weight-averaged molar mass was determined using a single ideal species model (see Methods). Mean value masses for the upper and lower plateaux were 138±6 kDa and 43±1 kDa respectively (averages of three measures). The molecular mass of native rEPclA (0 M GuHCl) is consistent with three times that of denatured rEPclA (see text). The transition midpoint concentration is 2.38±0.09 M GuHCl.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3368898&req=5

pone-0037872-g003: Analysis by analytical ultracentrifugation of the average molar mass of a sample of purified rEPclA as a function of increasing concentration of guanidinium chloride (GuHCl).Weight-averaged molar mass was determined using a single ideal species model (see Methods). Mean value masses for the upper and lower plateaux were 138±6 kDa and 43±1 kDa respectively (averages of three measures). The molecular mass of native rEPclA (0 M GuHCl) is consistent with three times that of denatured rEPclA (see text). The transition midpoint concentration is 2.38±0.09 M GuHCl.
Mentions: The quaternary structure of rEPclA was investigated by sedimentation equilibrium analytical ultracentrifugation (AUC) at increasing concentrations of guanidinium chloride (GuHCl) (Figure 3). The relative molar mass of rEPclA at 0 M GuHCl was 138±6 kDa, corresponding to the predicted molecular weight of a trimer of rEPclA molecules (3×47 kDa). As the concentration of GuHCl increased, a transition from trimer to monomer was observed and the relative molar mass of rEPclA at 5 M GuHCl was 43 kDa, which is consistent with the predicted molecular weight of the rEPclA monomer. Thus, rEPclA trimers dissociate into monomers as the GuHCl concentration increases; the trimer-to-monomer transition point was estimated at around 2.5 M GuHCl.

Bottom Line: Under the electron microscope, collagen-like proteins from E. coli O157:H7 show a dumbbell shape, with two globular domains joined by a hinged stalk.This morphology is consistent with their likely role as trimeric phage side-tail proteins that participate in the attachment of phage particles to E. coli target cells, either directly or through assembly with other phage tail proteins.Thus, collagen-like proteins in enterohaemorrhagic E. coli genomes may have a direct role in the dissemination of virulence-related genes through infection of harmless strains by induced bacteriophages.

View Article: PubMed Central - PubMed

Affiliation: Manchester Interdisciplinary Biocentre, University of Manchester, Manchester, United Kingdom.

ABSTRACT
The genome sequences of enterohaemorrhagic E. coli O157:H7 strains show multiple open-reading frames with collagen-like sequences that are absent from the common laboratory strain K-12. These putative collagens are included in prophages embedded in O157:H7 genomes. These prophages carry numerous genes related to strain virulence and have been shown to be inducible and capable of disseminating virulence factors by horizontal gene transfer. We have cloned two collagen-like proteins from E. coli O157:H7 into a laboratory strain and analysed the structure and conformation of the recombinant proteins and several of their constituting domains by a variety of spectroscopic, biophysical, and electron microscopy techniques. We show that these molecules exhibit many of the characteristics of vertebrate collagens, including trimer formation and the presence of a collagen triple helical domain. They also contain a C-terminal trimerization domain, and a trimeric α-helical coiled-coil domain with an unusual amino acid sequence almost completely lacking leucine, valine or isoleucine residues. Intriguingly, these molecules show high thermal stability, with the collagen domain being more stable than those of vertebrate fibrillar collagens, which are much longer and post-translationally modified. Under the electron microscope, collagen-like proteins from E. coli O157:H7 show a dumbbell shape, with two globular domains joined by a hinged stalk. This morphology is consistent with their likely role as trimeric phage side-tail proteins that participate in the attachment of phage particles to E. coli target cells, either directly or through assembly with other phage tail proteins. Thus, collagen-like proteins in enterohaemorrhagic E. coli genomes may have a direct role in the dissemination of virulence-related genes through infection of harmless strains by induced bacteriophages.

Show MeSH
Related in: MedlinePlus