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Contribution of membrane-binding and enzymatic domains of penicillin binding protein 5 to maintenance of uniform cellular morphology of Escherichia coli.
- Source :
-
Journal of bacteriology [J Bacteriol] 2002 Jul; Vol. 184 (13), pp. 3630-9. - Publication Year :
- 2002
-
Abstract
- Four low-molecular-weight penicillin binding proteins (LMW PBPs) of Escherichia coli are closely related and have similar DD-carboxypeptidase activities (PBPs 4, 5, and 6 and DacD). However, only one, PBP 5, has a demonstrated physiological function. In its absence, certain mutants of E. coli have altered diameters and lose their uniform outer contour, resulting in morphologically aberrant cells. To determine what differentiates the activities of these LMW PBPs, we constructed fusion proteins combining portions of PBP 5 with fragments of other DD-carboxypeptidases to see which hybrids restored normal morphology to a strain lacking PBP 5. Functional complementation occurred when truncated PBP 5 was combined with the terminal membrane anchor sequences of PBP 6 or DacD. However, complementation was not restored by the putative carboxy-terminal anchor of PBP 4 or by a transmembrane region of the osmosensor protein ProW, even though these hybrids were membrane bound. Site-directed mutagenesis of the carboxy terminus of PBP 5 indicated that complementation required a generalized amphipathic membrane anchor but that no specific residues in this region seemed to be required. A functional fusion protein was produced by combining the N-terminal enzymatic domain of PBP 5 with the C-terminal beta-sheet domain of PBP 6. In contrast, the opposite hybrid of PBP 6 to PBP 5 was not functional. The results suggest that the mode of PBP 5 membrane anchoring is important, that the mechanism entails more than a simple mechanical tethering of the enzyme to the outer face of the inner membrane, and that the physiological differences among the LMW PBPs arise from structural differences in the DD-carboxypeptidase enzymatic core.
- Subjects :
- Amino Acid Sequence
Base Sequence
Binding Sites
Carrier Proteins genetics
Endopeptidases metabolism
Molecular Sequence Data
Muramoylpentapeptide Carboxypeptidase genetics
Mutagenesis, Site-Directed
Penicillin-Binding Proteins
Polymerase Chain Reaction methods
Protein Structure, Tertiary
Recombinant Fusion Proteins genetics
Recombinant Fusion Proteins metabolism
Sequence Homology, Amino Acid
Bacterial Proteins
Carrier Proteins metabolism
Cell Membrane metabolism
Dipeptidases
Escherichia coli physiology
Hexosyltransferases
Muramoylpentapeptide Carboxypeptidase metabolism
Peptidyl Transferases
Subjects
Details
- Language :
- English
- ISSN :
- 0021-9193
- Volume :
- 184
- Issue :
- 13
- Database :
- MEDLINE
- Journal :
- Journal of bacteriology
- Publication Type :
- Academic Journal
- Accession number :
- 12057958
- Full Text :
- https://doi.org/10.1128/JB.184.13.3630-3639.2002