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The essential peptidoglycan glycosyltransferase MurG forms a complex with proteins involved in lateral envelope growth as well as with proteins involved in cell division in Escherichia coli.

Mohammadi T, Karczmarek A, Crouvoisier M, Bouhss A, Mengin-Lecreulx D, den Blaauwen T - Mol. Microbiol. (2007)

Bottom Line: In view of this, the loss of rod shape of DeltamreBCD strain could be ascribed to the loss of MurG membrane localization.Consequently, this could prevent the localized supply of the lipid II precursor to the peptidoglycan synthesizing machinery involved in cell elongation.A model representing the first complex is proposed.

View Article: PubMed Central - PubMed

Affiliation: Molecular Cytology, Swammerdam Institute for Life Sciences, University of Amsterdam, Kruislaan 316, 1098 SM Amsterdam, PO Box 194062, 1090 GB Amsterdam, The Netherlands.

ABSTRACT
In Escherichia coli many enzymes including MurG are directly involved in the synthesis and assembly of peptidoglycan. MurG is an essential glycosyltransferase catalysing the last intracellular step of peptidoglycan synthesis. To elucidate its role during elongation and division events, localization of MurG using immunofluorescence microscopy was performed. MurG exhibited a random distribution in the cell envelope with a relatively higher intensity at the division site. This mid-cell localization was dependent on the presence of a mature divisome. Its localization in the lateral cell wall appeared to require the presence of MreCD. This could be indicative of a potential interaction between MurG and other proteins. Investigating this by immunoprecipitation revealed the association of MurG with MreB and MraY in the same protein complex. In view of this, the loss of rod shape of DeltamreBCD strain could be ascribed to the loss of MurG membrane localization. Consequently, this could prevent the localized supply of the lipid II precursor to the peptidoglycan synthesizing machinery involved in cell elongation. It is postulated that the involvement of MurG in the peptidoglycan synthesis concurs with two complexes, one implicated in cell elongation and the other in division. A model representing the first complex is proposed.

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Fluorescence intensity profiles of LMC509 (FtsZ(Ts), A and B), LMC510 (PBP3(Ts), C), LMC531 (FtsQ(Ts), D) and LMC500 (wild-type, E) cells. After growth at 28°C in 1/2 GB1 LMC509 cells were shifted to 42°C for 2 MDs, and immunolabelled with anti-MurG (A, 28°C) and (B, 42°C). LMC510 and LMC531 cells were grown at 28°C in GB1 to steady state and shifted to 42°C for 2 MDs, and immunolabelled with anti-MurG. LMC500 cells were grown at 28°C in GB1 after which aztreonam was added and growth was continued for 2 MDs before the cells were immunolabelled with anti-MurG (E). Similar patterns shown in A were also obtained for the other temperature-sensitive strains when grown at 28°C in GB1. The average normalized cell length is given on the x-axis. The y-axis represents the average normalized fluorescence intensity.
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fig03: Fluorescence intensity profiles of LMC509 (FtsZ(Ts), A and B), LMC510 (PBP3(Ts), C), LMC531 (FtsQ(Ts), D) and LMC500 (wild-type, E) cells. After growth at 28°C in 1/2 GB1 LMC509 cells were shifted to 42°C for 2 MDs, and immunolabelled with anti-MurG (A, 28°C) and (B, 42°C). LMC510 and LMC531 cells were grown at 28°C in GB1 to steady state and shifted to 42°C for 2 MDs, and immunolabelled with anti-MurG. LMC500 cells were grown at 28°C in GB1 after which aztreonam was added and growth was continued for 2 MDs before the cells were immunolabelled with anti-MurG (E). Similar patterns shown in A were also obtained for the other temperature-sensitive strains when grown at 28°C in GB1. The average normalized cell length is given on the x-axis. The y-axis represents the average normalized fluorescence intensity.

Mentions: To investigate whether the presence of MurG at mid-cell was division dependent, its localization was studied in the strain LMC509 that expresses the temperature-sensitive FtsZ84(Ts) protein, which is not able to polymerize at mid-cell and causes filamentation at the restrictive temperature (Addinall et al., 1997). MurG showed normal mid-cell localization in this strain at 28°C (Fig. 3A). However, at 42°C MurG foci were randomly distributed in the cell envelope (Fig. 3B). This suggests that the mid-cell localization of MurG is related to cell division. The earliest event in cell division is the assembly of the FtsZ-ring. This is followed by the assembly of a number of other cell division proteins at mid-cell about 1/5 of a generation time later just before the onset of the constriction process (Aarsman et al., 2005). To examine whether the MurG mid-cell localization is dependent on an early or a late event in the process of cell division, localization studies of MurG were further performed in strain LMC510, which contains the ftsI2158 that expresses a PBP3(Ts) protein. PBP3 is involved in peptidoglycan synthesis and localizes to the division site during the later stages of cell growth and throughout septation (Weiss et al., 1999; Aarsman et al., 2005). At 42°C the PBP3(Ts) filaments showed blunt constriction where mid-cell localization of MurG was not observed (Fig. 3C and Fig. S4A). This implies that the position of MurG is dependent on the presence of PBP3.


The essential peptidoglycan glycosyltransferase MurG forms a complex with proteins involved in lateral envelope growth as well as with proteins involved in cell division in Escherichia coli.

Mohammadi T, Karczmarek A, Crouvoisier M, Bouhss A, Mengin-Lecreulx D, den Blaauwen T - Mol. Microbiol. (2007)

Fluorescence intensity profiles of LMC509 (FtsZ(Ts), A and B), LMC510 (PBP3(Ts), C), LMC531 (FtsQ(Ts), D) and LMC500 (wild-type, E) cells. After growth at 28°C in 1/2 GB1 LMC509 cells were shifted to 42°C for 2 MDs, and immunolabelled with anti-MurG (A, 28°C) and (B, 42°C). LMC510 and LMC531 cells were grown at 28°C in GB1 to steady state and shifted to 42°C for 2 MDs, and immunolabelled with anti-MurG. LMC500 cells were grown at 28°C in GB1 after which aztreonam was added and growth was continued for 2 MDs before the cells were immunolabelled with anti-MurG (E). Similar patterns shown in A were also obtained for the other temperature-sensitive strains when grown at 28°C in GB1. The average normalized cell length is given on the x-axis. The y-axis represents the average normalized fluorescence intensity.
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Related In: Results  -  Collection

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fig03: Fluorescence intensity profiles of LMC509 (FtsZ(Ts), A and B), LMC510 (PBP3(Ts), C), LMC531 (FtsQ(Ts), D) and LMC500 (wild-type, E) cells. After growth at 28°C in 1/2 GB1 LMC509 cells were shifted to 42°C for 2 MDs, and immunolabelled with anti-MurG (A, 28°C) and (B, 42°C). LMC510 and LMC531 cells were grown at 28°C in GB1 to steady state and shifted to 42°C for 2 MDs, and immunolabelled with anti-MurG. LMC500 cells were grown at 28°C in GB1 after which aztreonam was added and growth was continued for 2 MDs before the cells were immunolabelled with anti-MurG (E). Similar patterns shown in A were also obtained for the other temperature-sensitive strains when grown at 28°C in GB1. The average normalized cell length is given on the x-axis. The y-axis represents the average normalized fluorescence intensity.
Mentions: To investigate whether the presence of MurG at mid-cell was division dependent, its localization was studied in the strain LMC509 that expresses the temperature-sensitive FtsZ84(Ts) protein, which is not able to polymerize at mid-cell and causes filamentation at the restrictive temperature (Addinall et al., 1997). MurG showed normal mid-cell localization in this strain at 28°C (Fig. 3A). However, at 42°C MurG foci were randomly distributed in the cell envelope (Fig. 3B). This suggests that the mid-cell localization of MurG is related to cell division. The earliest event in cell division is the assembly of the FtsZ-ring. This is followed by the assembly of a number of other cell division proteins at mid-cell about 1/5 of a generation time later just before the onset of the constriction process (Aarsman et al., 2005). To examine whether the MurG mid-cell localization is dependent on an early or a late event in the process of cell division, localization studies of MurG were further performed in strain LMC510, which contains the ftsI2158 that expresses a PBP3(Ts) protein. PBP3 is involved in peptidoglycan synthesis and localizes to the division site during the later stages of cell growth and throughout septation (Weiss et al., 1999; Aarsman et al., 2005). At 42°C the PBP3(Ts) filaments showed blunt constriction where mid-cell localization of MurG was not observed (Fig. 3C and Fig. S4A). This implies that the position of MurG is dependent on the presence of PBP3.

Bottom Line: In view of this, the loss of rod shape of DeltamreBCD strain could be ascribed to the loss of MurG membrane localization.Consequently, this could prevent the localized supply of the lipid II precursor to the peptidoglycan synthesizing machinery involved in cell elongation.A model representing the first complex is proposed.

View Article: PubMed Central - PubMed

Affiliation: Molecular Cytology, Swammerdam Institute for Life Sciences, University of Amsterdam, Kruislaan 316, 1098 SM Amsterdam, PO Box 194062, 1090 GB Amsterdam, The Netherlands.

ABSTRACT
In Escherichia coli many enzymes including MurG are directly involved in the synthesis and assembly of peptidoglycan. MurG is an essential glycosyltransferase catalysing the last intracellular step of peptidoglycan synthesis. To elucidate its role during elongation and division events, localization of MurG using immunofluorescence microscopy was performed. MurG exhibited a random distribution in the cell envelope with a relatively higher intensity at the division site. This mid-cell localization was dependent on the presence of a mature divisome. Its localization in the lateral cell wall appeared to require the presence of MreCD. This could be indicative of a potential interaction between MurG and other proteins. Investigating this by immunoprecipitation revealed the association of MurG with MreB and MraY in the same protein complex. In view of this, the loss of rod shape of DeltamreBCD strain could be ascribed to the loss of MurG membrane localization. Consequently, this could prevent the localized supply of the lipid II precursor to the peptidoglycan synthesizing machinery involved in cell elongation. It is postulated that the involvement of MurG in the peptidoglycan synthesis concurs with two complexes, one implicated in cell elongation and the other in division. A model representing the first complex is proposed.

Show MeSH
Related in: MedlinePlus