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A Staphylococcus aureus ypfP mutant with strongly reduced lipoteichoic acid (LTA) content: LTA governs bacterial surface properties and autolysin activity.

Fedtke I, Mader D, Kohler T, Moll H, Nicholson G, Biswas R, Henseler K, Götz F, Zähringer U, Peschel A - Mol. Microbiol. (2007)

Bottom Line: The ypfP gene responsible for biosynthesis of a glycolipid found in LTA was deleted in Staphylococcus aureus SA113, causing 87% reduction of the LTA content.However, the autolytic activity of the mutant was strongly reduced demonstrating a role of LTA in controlling autolysin activity.Moreover, the hydrophobicity of the LTA mutant was altered and its ability to form biofilms on plastic was completely abrogated indicating a profound impact of LTA on physicochemical properties of bacterial surfaces.

View Article: PubMed Central - PubMed

Affiliation: Cellular and Molecular Microbiology Division, University of Tübingen, Department of Medical Microbiology and Hygiene, 72076 Tübingen, Germany.

ABSTRACT
Many Gram-positive bacteria produce lipoteichoic acid (LTA) polymers whose physiological roles have remained a matter of debate because of the lack of LTA-deficient mutants. The ypfP gene responsible for biosynthesis of a glycolipid found in LTA was deleted in Staphylococcus aureus SA113, causing 87% reduction of the LTA content. Mass spectrometry and nuclear magnetic resonance spectroscopy revealed that the mutant LTA contained a diacylglycerol anchor instead of the glycolipid, whereas the remaining part was similar to the wild-type polymer except that it was shorter. The LTA mutant strain revealed no major changes in patterns of cell wall proteins or autolytic enzymes compared with the parental strain indicating that LTA may be less important in S. aureus protein attachment than previously thought. However, the autolytic activity of the mutant was strongly reduced demonstrating a role of LTA in controlling autolysin activity. Moreover, the hydrophobicity of the LTA mutant was altered and its ability to form biofilms on plastic was completely abrogated indicating a profound impact of LTA on physicochemical properties of bacterial surfaces. We propose to consider LTA and its biosynthetic enzymes as targets for new antibiofilm strategies.

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Related in: MedlinePlus

GLC-MS spectra of the per-O-acetylated glycan isolated from S. aureus SA113 wild-type LTA. The EI mass spectrum (top) shows the predominant m/z = 159 fragment for the Gro unit. Other characteristic fragments originating from the terminal hexose in the glycerol gentiobiose pseudotrisaccharide are shown with enlarged intensity (5×) on the right. The base peak (m/z = 812) in the CI MS spectrum (bottom) of the per-O-acetylated glycan represents the pseudomolecular ion [M+NH4]+ of the per-O-acetylated glycerol β-gentiobiose pseudotrisaccharide (Glc-O-Glc-O-Gro).
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fig03: GLC-MS spectra of the per-O-acetylated glycan isolated from S. aureus SA113 wild-type LTA. The EI mass spectrum (top) shows the predominant m/z = 159 fragment for the Gro unit. Other characteristic fragments originating from the terminal hexose in the glycerol gentiobiose pseudotrisaccharide are shown with enlarged intensity (5×) on the right. The base peak (m/z = 812) in the CI MS spectrum (bottom) of the per-O-acetylated glycan represents the pseudomolecular ion [M+NH4]+ of the per-O-acetylated glycerol β-gentiobiose pseudotrisaccharide (Glc-O-Glc-O-Gro).

Mentions: Gas-liquid chromatography (GLC)-MS analyses of the fatty acids in the LTA of both strains revealed an identical fatty acid profile (iso-15:0 and iso-17:0) whereby iso-15:0 dominated. Methanolysis followed by per-O-acetylation and subsequent GLC-MS revealed the expected components of the repeating glycerophosphate in both LTAs, whereby unsubstituted Gro, Gro phosphate di-methylester (Gro-P), alanylated Gro (Gro-d-Ala) and 2-acetamido-2-deoxy-α-d-glucopyranosyl-substituted Gro (Gro-GlcNAc) were found in comparable amounts and proportions in LTA preparations from parental and mutant strains. However, only in case of the wild-type strain, a late eluting peak (46.46 min) could be detected in the GLC-MS (data not shown) suggesting that this compound, maybe a short oligosaccharide, is not present in the mutant. The electron impact (EI) mass spectrum (Fig. 3) showed diagnostic fragment ions at m/z = 331 for a terminal peracetylated hexosyl (GlcII) and m/z = 159 assigned to the Gro residue. Chemical ionization (CI) with ammonia revealed a pseudomolecular ion ([M + NH4]+m/z = 812), which was compatible with the per-O-acetylated glyceryl β-gentiobioside (Glc-O-Glc-O-Gro) (Fig. 3).


A Staphylococcus aureus ypfP mutant with strongly reduced lipoteichoic acid (LTA) content: LTA governs bacterial surface properties and autolysin activity.

Fedtke I, Mader D, Kohler T, Moll H, Nicholson G, Biswas R, Henseler K, Götz F, Zähringer U, Peschel A - Mol. Microbiol. (2007)

GLC-MS spectra of the per-O-acetylated glycan isolated from S. aureus SA113 wild-type LTA. The EI mass spectrum (top) shows the predominant m/z = 159 fragment for the Gro unit. Other characteristic fragments originating from the terminal hexose in the glycerol gentiobiose pseudotrisaccharide are shown with enlarged intensity (5×) on the right. The base peak (m/z = 812) in the CI MS spectrum (bottom) of the per-O-acetylated glycan represents the pseudomolecular ion [M+NH4]+ of the per-O-acetylated glycerol β-gentiobiose pseudotrisaccharide (Glc-O-Glc-O-Gro).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2169524&req=5

fig03: GLC-MS spectra of the per-O-acetylated glycan isolated from S. aureus SA113 wild-type LTA. The EI mass spectrum (top) shows the predominant m/z = 159 fragment for the Gro unit. Other characteristic fragments originating from the terminal hexose in the glycerol gentiobiose pseudotrisaccharide are shown with enlarged intensity (5×) on the right. The base peak (m/z = 812) in the CI MS spectrum (bottom) of the per-O-acetylated glycan represents the pseudomolecular ion [M+NH4]+ of the per-O-acetylated glycerol β-gentiobiose pseudotrisaccharide (Glc-O-Glc-O-Gro).
Mentions: Gas-liquid chromatography (GLC)-MS analyses of the fatty acids in the LTA of both strains revealed an identical fatty acid profile (iso-15:0 and iso-17:0) whereby iso-15:0 dominated. Methanolysis followed by per-O-acetylation and subsequent GLC-MS revealed the expected components of the repeating glycerophosphate in both LTAs, whereby unsubstituted Gro, Gro phosphate di-methylester (Gro-P), alanylated Gro (Gro-d-Ala) and 2-acetamido-2-deoxy-α-d-glucopyranosyl-substituted Gro (Gro-GlcNAc) were found in comparable amounts and proportions in LTA preparations from parental and mutant strains. However, only in case of the wild-type strain, a late eluting peak (46.46 min) could be detected in the GLC-MS (data not shown) suggesting that this compound, maybe a short oligosaccharide, is not present in the mutant. The electron impact (EI) mass spectrum (Fig. 3) showed diagnostic fragment ions at m/z = 331 for a terminal peracetylated hexosyl (GlcII) and m/z = 159 assigned to the Gro residue. Chemical ionization (CI) with ammonia revealed a pseudomolecular ion ([M + NH4]+m/z = 812), which was compatible with the per-O-acetylated glyceryl β-gentiobioside (Glc-O-Glc-O-Gro) (Fig. 3).

Bottom Line: The ypfP gene responsible for biosynthesis of a glycolipid found in LTA was deleted in Staphylococcus aureus SA113, causing 87% reduction of the LTA content.However, the autolytic activity of the mutant was strongly reduced demonstrating a role of LTA in controlling autolysin activity.Moreover, the hydrophobicity of the LTA mutant was altered and its ability to form biofilms on plastic was completely abrogated indicating a profound impact of LTA on physicochemical properties of bacterial surfaces.

View Article: PubMed Central - PubMed

Affiliation: Cellular and Molecular Microbiology Division, University of Tübingen, Department of Medical Microbiology and Hygiene, 72076 Tübingen, Germany.

ABSTRACT
Many Gram-positive bacteria produce lipoteichoic acid (LTA) polymers whose physiological roles have remained a matter of debate because of the lack of LTA-deficient mutants. The ypfP gene responsible for biosynthesis of a glycolipid found in LTA was deleted in Staphylococcus aureus SA113, causing 87% reduction of the LTA content. Mass spectrometry and nuclear magnetic resonance spectroscopy revealed that the mutant LTA contained a diacylglycerol anchor instead of the glycolipid, whereas the remaining part was similar to the wild-type polymer except that it was shorter. The LTA mutant strain revealed no major changes in patterns of cell wall proteins or autolytic enzymes compared with the parental strain indicating that LTA may be less important in S. aureus protein attachment than previously thought. However, the autolytic activity of the mutant was strongly reduced demonstrating a role of LTA in controlling autolysin activity. Moreover, the hydrophobicity of the LTA mutant was altered and its ability to form biofilms on plastic was completely abrogated indicating a profound impact of LTA on physicochemical properties of bacterial surfaces. We propose to consider LTA and its biosynthetic enzymes as targets for new antibiofilm strategies.

Show MeSH
Related in: MedlinePlus