In vitro characterization of peptidoglycan-associated lipoprotein (PAL)-peptidoglycan and PAL-TolB interactions.
The Tol-peptidoglycan-associated lipoprotein (PAL) system of Escherichia coli is a multiprotein complex of the envelope involved in maintaining outer membrane integrity. PAL and the periplasmic protein TolB, two components of this complex, are interacting with each other, and they have also been reported to interact with OmpA and the major lipoprotein, two proteins interacting with the peptidoglycan. All these interactions suggest a role of the Tol-PAL system in anchoring the outer membrane to the peptidoglycan. Therefore, we were interested in better understanding the interaction between PAL and the peptidoglycan. We designed an in vitro interaction assay based on the property of purified peptidoglycan to be pelleted by ultracentrifugation. Using this assay, we showed that a purified PAL protein interacted in vitro with pure peptidoglycan. A peptide competition experiment further demonstrated that the region from residues 89 to 130 of PAL was sufficient to bind the peptidoglycan. Moreover, the fact that this same region of PAL was also binding to TolB suggested that these two interactions were exclusive. Indeed, the TolB-PAL complex appeared not to be associated with the peptidoglycan. This led us to the conclusion that PAL may exist in two forms in the cell envelope, one bound to TolB and the other bound to the peptidoglycan.
Bouveret
E
E
Laboratoire d'Ingéniérie des Systèmes Macromoléculaires, Institut de Biologie Structurale et Microbiologie, Centre National de la Recherche Scientifique (CNRS), 13402 Marseille Cedex 20, France.
Bénédetti
H
H
Rigal
A
A
Loret
E
E
Lazdunski
C
C
D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins / Q000378:metabolism; D001426:Bacterial Proteins / Q000378:metabolism*; D001665:Binding Sites; D001667:Binding, Competitive; D002462:Cell Membrane / Q000378:metabolism; D002473:Cell Wall / Q000378:metabolism; D003432:Cross-Linking Reagents; D004926:Escherichia coli; D029968:Escherichia coli Proteins; D006193:Haemophilus influenzae; D008074:Lipoproteins / Q000302:isolation & purification / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000378:metabolism; D010457:Peptidoglycan / Q000302:isolation & purification / Q000378:metabolism*; D019897:Periplasm / Q000378:metabolism; D033903:Periplasmic Proteins; D011485:Protein Binding; D011509:Proteoglycans
D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D003432:Cross-Linking Reagents; D029968:Escherichia coli Proteins; C031163:ExcC protein, E coli; D008074:Lipoproteins; D008565:Membrane Proteins; D010446:Peptide Fragments; D010457:Peptidoglycan; D033903:Periplasmic Proteins; D011509:Proteoglycans; C062094:tolB protein, E coli; C069988:PplA protein, Legionella pneumophila