Influence of Pilin Glycosylation on
            <i>Pseudomonas aeruginosa</i>
            1244 Pilus Function

Smedley, James G.; Jewell, Erica; Roguskie, Jennifer; Horzempa, Joseph; Syboldt, Andrew; Stolz, Donna B.; Castric, Peter

doi:10.1128/iai.73.12.7922-7931.2005

Cited by 106 publications

(100 citation statements)

References 53 publications

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“…The only MglAregulated gene identified in these studies that is annotated to potentially play a role in Tfp biogenesis corresponds to FTN_0431 (FTT0905 in Schu S4, FTL0425 in LVS). The encoded protein shares homology with the Tfp glycosylation protein PilO from Pseudomonas aeruginosa (Smedley et al, 2005). Transcript levels for FTN_0431 were lower in the mglA mutant than in the wild-type strain, confirming that MglA regulates FTN_0431 transcription (Fig.…”

Section: Mgla Regulates Tfp Expressionsupporting

confidence: 48%

“…holarctica LVS both identified an MglA-regulated gene annotated as a Tfp glycosylation protein (FTT0905; (Brotcke et al, 2006;Charity et al, 2007) with homology to the Tfp glycosylation protein PilO of P. aeruginosa (25 % identity, 45 % similarity; Kus et al, 2004). Although pilO is not required for P. aeruginosa pilus expression, evidence suggests that glycosylated pili enhance its virulence (Smedley et al, 2005). However, PilO homologues have been shown to be important for pilus expression in Synechocystis (Yoshihara et al, 2001) and Pseudomonas syringae (Roine et al, 1998).…”

Section: Mgla Regulation Of Tfp Expression In F Tularensismentioning

confidence: 99%

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Characterization of the Francisella tularensis subsp. novicida type IV pilus

et al. 2008

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Francisella tularensis causes the disease tularaemia. Type IV pili (Tfp) genes are present in the genomes of all F. tularensis subspecies. We show that the wild-type F. tularensis subsp. novicida expresses pilus fibres on its surface, and mutations in the Tfp genes pilF and pilT disrupt pilus biogenesis. Mutations in other Tfp genes (pilQ and pilG) do not eliminate pilus expression. A mutation in pilE4 eliminates pilus expression, whereas mutations in the other pilin subunits pilE1-3 and pilE5 do not, suggesting that pilE4 is the major pilus structural subunit. The virulence regulator MglA is required for pilus expression, and it regulates the transcription of a putative Tfp glycosylation gene (FTN0431). However, MglA does not regulate transcription of pilF, pilT or pilE4, and a strain lacking FTN0431 still expresses pili; thus, it is unclear how MglA regulates pilus expression. Only pilF was also required for protein secretion, while pilE4 and pilT were not, indicating that there is very little overlap of the protein secretion/Tfp functions of the pil genes. The protein secretion component pilE1 was more important for in vitro intramacrophage growth and mouse virulence than the Tfp component pilE4. Our results provide the first genetic characterization of the novel Tfp system of F. tularensis. INTRODUCTIONFrancisella tularensis is a highly infectious bacterium that causes tularaemia. F. tularensis is found in many different animal hosts and can be transmitted by arthropod vectors (Ellis et al., 2002). Humans can acquire the bacteria by a number of different routes, but inhalation of low doses of the organism can lead to a serious pneumonic form of disease that has a high mortality rate, and this has led to the classification of this bacterium as a category A biothreat agent (Dennis et al., 2001). F. tularensis is further divided into different subspecies (Oyston et al., 2004). F. tularensis subsp. tularensis has historically been associated with bioweapons development, and is reported to have the highest virulence for humans (McLendon et al., 2006). F. tularensis subsp. holarctica is also infectious in humans but typically with a less severe outcome (Sjostedt, 2003). An attenuated 'live vaccine strain' (LVS) was derived by repeated passage of subsp. holarctica in the laboratory; despite the unclear nature of the attenuating mutation(s) in this strain, it is frequently used as a laboratory model for tularaemia (Anthony & Kongshavn, 1987;Eigelsbach et al., 1951). F. tularensis subsp. novicida has low virulence for humans, but maintains high virulence in mice (Kieffer et al., 2003;Lauriano et al., 2004). Whole-genome sequencing has revealed that all three subspecies are closely related, with the less virulent (for humans) subsp. novicida showing less genomic decay than the more virulent subsp. holarctica and subsp. tularensis (Larsson et al., 2005;Petrosino et al., 2006;Rohmer et al., 2006).The ability of F. tularensis to survive and replicate within macrophages has been linked to its virulence (Anthony et al., 19...

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Section: Mgla Regulates Tfp Expressionsupporting

confidence: 48%

Section: Mgla Regulation Of Tfp Expression In F Tularensismentioning

confidence: 99%

Characterization of the Francisella tularensis subsp. novicida type IV pilus

et al. 2008

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“…Each PilA subunit of the pilus fiber is decorated with a single trisaccharide (386). The glycan is evenly distributed over the fiber surface, at the seam between two adjacent pilins (285). The trisaccharide consists of 5-N-␤-OHC 4 7NfmPse, xylose, and FucNAc (386) linked to C-terminal serine 148 (387).…”

Section: Protein Glycosylationmentioning

confidence: 99%

“…Glycosylation of proteins in bacteria has been shown to modulate the physicochemical properties of their substrates. Glycan modifications of proteins can protect them against proteolysis (e.g., AIDA-I of E. coli 2787 [282]), confer extra stability (e.g., C. jejuni proteins [283]), enhance activity (e.g., gingipains of Porphyromonas gingivalis [284]), and change the surface properties (e.g., pili of P. aeruginosa 1244 [285]). In addition, glycoproteins play a role in interactions between bacteria and their surroundings.…”

Section: Protein Glycosylationmentioning

confidence: 99%

The Sweet Tooth of Bacteria: Common Themes in Bacterial Glycoconjugates

Tytgat

Lebeer

2014

Microbiol Mol Biol Rev

129

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SUMMARY Humans have been increasingly recognized as being superorganisms, living in close contact with a microbiota on all their mucosal surfaces. However, most studies on the human microbiota have focused on gaining comprehensive insights into the composition of the microbiota under different health conditions (e.g., enterotypes), while there is also a need for detailed knowledge of the different molecules that mediate interactions with the host. Glycoconjugates are an interesting class of molecules for detailed studies, as they form a strain-specific barcode on the surface of bacteria, mediating specific interactions with the host. Strikingly, most glycoconjugates are synthesized by similar biosynthesis mechanisms. Bacteria can produce their major glycoconjugates by using a sequential or an en bloc mechanism, with both mechanistic options coexisting in many species for different macromolecules. In this review, these common themes are conceptualized and illustrated for all major classes of known bacterial glycoconjugates, with a special focus on the rather recently emergent field of glycosylated proteins. We describe the biosynthesis and importance of glycoconjugates in both pathogenic and beneficial bacteria and in both Gram-positive and -negative organisms. The focus lies on microorganisms important for human physiology. In addition, the potential for a better knowledge of bacterial glycoconjugates in the emerging field of glycoengineering and other perspectives is discussed.

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“…2), a normally nonglycosylated pilin (44), with those of 1244 pilin and assayed for glycosylation. When plasmid pSD5, which contained the pilA gene from PA103 behind a tac promoter, was expressed in strain 1244, PA103 pilin produced had the same apparent molecular weight as that of wild type (Fig.…”

Section: Structural Requirements Of the Pilin Glycosylationmentioning

confidence: 99%

Glycosylation Substrate Specificity of Pseudomonas aeruginosa 1244 Pilin

Horzempa

Comer

Davis

et al. 2006

Journal of Biological Chemistry

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The ␤-carbon of the Pseudomonas aeruginosa 1244 pilin C-terminal Ser is a site of glycosylation. The present study was conducted to determine the pilin structures necessary for glycosylation. It was found that although Thr could be tolerated at the pilin C terminus, the blocking of the Ser carboxyl group with the addition of an Ala prevented glycosylation. Pilin from strain PA103 was not glycosylated by P. aeruginosa 1244, even when the C-terminal residue was converted to Ser. Substituting the disulfide loop region of strain PA103 pilin with that of strain 1244 allowed glycosylation to take place. Neither conversion of 1244 pilin disulfide loop Cys residues to Ala nor the deletion of segments of this structure prevented glycosylation. It was noted that the PA103 pilin disulfide loop environment was electronegative, whereas that of strain 1244 pilin had an overall positive charge. Insertion of a positive charge into the PA103 pilin disulfide loop of a mutant containing Ser at the C terminus allowed glycosylation to take place. Extending the "tail" region of the PA103 mutant pilin containing Ser at its terminus resulted in robust glycosylation. These results suggest that the terminal Ser is the major pilin glycosylation recognition feature and that this residue cannot be substituted at its carboxyl group. Although no other specific recognition features are present, the pilin surface must be compatible with the reaction apparatus for glycosylation to occur.Pseudomonas aeruginosa is a Gram-negative bacterium that expresses polar filaments called type IV pili. These pili are polymers of a primarily proteinaceous subunit referred to as pilin. Although all known type IV pilins are modified by the cleavage of a leader sequence followed by the N-methylation of the exposed Phe (1), the pilin of P. aeruginosa 1244 is additionally glycosylated. This glycosylation requires the presence of PilO, an enzyme whose coding sequence is in the same operon as pilA, the pilin structural gene (2).The first discovered examples of prokaryotic glycoproteins were archaeal S-layer proteins (3). Since this finding, numerous examples of protein glycosylation in eubacteria have been identified (4 -8), indicating that this protein modification is distributed among all of the biological kingdoms. Proteins destined for glycosylation may be modified at a single or multiple sites along the peptide chain (7,8) in which the glycan may be one sugar or an assortment of diverse saccharides. Oligosaccharide assembly can take place either by sequential sugar addition to the target protein or by synthesis prior to glycosylation in which the entire pre-formed glycan is transferred as a single unit (7,8). Numerous protein/sugar linkages have been characterized involving many functional groups present on peptide chains and most of the commonly encountered monosaccharide moieties (8), in addition to exotic prokaryotic sugars such as bacillosamine, pseudaminic acid, and derivatives (7). Carbohydrates in eukaryotes and prokaryotes, alike, are usually bound to th...

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Influence of Pilin Glycosylation on Pseudomonas aeruginosa 1244 Pilus Function

Cited by 106 publications

References 53 publications

Characterization of the Francisella tularensis subsp. novicida type IV pilus

Characterization of the Francisella tularensis subsp. novicida type IV pilus

The Sweet Tooth of Bacteria: Common Themes in Bacterial Glycoconjugates

Glycosylation Substrate Specificity of Pseudomonas aeruginosa 1244 Pilin

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