Abstract
The construction of hybrid proteins of PBP1B and PBP3 has been described. One hybrid protein (PBP1B/3) contained the transglycosylase domain of PBP1B and the transpeptidase domain of PBP3. In the other hybrid protein, the putative transglycosylase domain of PBP3 was coupled to the transpeptidase domain of PBP1B (PBP3/1B). The hybrid proteins were localized in the cell envelope in a similar way as the wild-type PBP1B. In vitro isolates of the strains containing the hybrid proteins had a transglycosylase activity intermediate between that of wild-type PBP1B-producing strain and that of a PBP1B overproducer. Analysis with specific antibiotics against PBP1A/1B and PBP3 and mutant analysis in strains containing PBP3/1B revealed no detectable effects in vivo compared with wild-type strains. The same was shown for PBP1B/3 when the experiments were performed in a recA background. The data indicate that the hybrid proteins cannot replace native penicillin-binding proteins. This finding suggests that functional high-molecular-weight penicillin-binding protein specificity is at least in part determined by the unique combination of the two functional domains.
Full Text
The Full Text of this article is available as a PDF (256.0 KB).
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Broome-Smith J. K., Edelman A., Yousif S., Spratt B. G. The nucleotide sequences of the ponA and ponB genes encoding penicillin-binding protein 1A and 1B of Escherichia coli K12. Eur J Biochem. 1985 Mar 1;147(2):437–446. doi: 10.1111/j.1432-1033.1985.tb08768.x. [DOI] [PubMed] [Google Scholar]
- Casadaban M. J. Transposition and fusion of the lac genes to selected promoters in Escherichia coli using bacteriophage lambda and Mu. J Mol Biol. 1976 Jul 5;104(3):541–555. doi: 10.1016/0022-2836(76)90119-4. [DOI] [PubMed] [Google Scholar]
- Den Blaauwen T., Wientjes F. B., Kolk A. H., Spratt B. G., Nanninga N. Preparation and characterization of monoclonal antibodies against native membrane-bound penicillin-binding protein 1B of Escherichia coli. J Bacteriol. 1989 Mar;171(3):1394–1401. doi: 10.1128/jb.171.3.1394-1401.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Filip C., Fletcher G., Wulff J. L., Earhart C. F. Solubilization of the cytoplasmic membrane of Escherichia coli by the ionic detergent sodium-lauryl sarcosinate. J Bacteriol. 1973 Sep;115(3):717–722. doi: 10.1128/jb.115.3.717-722.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hedge P. J., Spratt B. G. Amino acid substitutions that reduce the affinity of penicillin-binding protein 3 of Escherichia coli for cephalexin. Eur J Biochem. 1985 Aug 15;151(1):111–121. doi: 10.1111/j.1432-1033.1985.tb09075.x. [DOI] [PubMed] [Google Scholar]
- Joris B., Ghuysen J. M., Dive G., Renard A., Dideberg O., Charlier P., Frère J. M., Kelly J. A., Boyington J. C., Moews P. C. The active-site-serine penicillin-recognizing enzymes as members of the Streptomyces R61 DD-peptidase family. Biochem J. 1988 Mar 1;250(2):313–324. doi: 10.1042/bj2500313. [DOI] [PMC free article] [PubMed] [Google Scholar]
- LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
- Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
- Nicholas R. A., Strominger J. L., Suzuki H., Hirota Y. Identification of the active site in penicillin-binding protein 3 of Escherichia coli. J Bacteriol. 1985 Oct;164(1):456–460. doi: 10.1128/jb.164.1.456-460.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Nicholas R. A., Suzuki H., Hirota Y., Strominger J. L. Purification and sequencing of the active site tryptic peptide from penicillin-binding protein 1b of Escherichia coli. Biochemistry. 1985 Jul 2;24(14):3448–3453. doi: 10.1021/bi00335a009. [DOI] [PubMed] [Google Scholar]
- Spratt B. G. Properties of the penicillin-binding proteins of Escherichia coli K12,. Eur J Biochem. 1977 Jan;72(2):341–352. doi: 10.1111/j.1432-1033.1977.tb11258.x. [DOI] [PubMed] [Google Scholar]
- Suzuki H., van Heijenoort Y., Tamura T., Mizoguchi J., Hirota Y., van Heijenoort J. In vitro peptidoglycan polymerization catalysed by penicillin binding protein 1b of Escherichia coli K-12. FEBS Lett. 1980 Feb 11;110(2):245–249. doi: 10.1016/0014-5793(80)80083-4. [DOI] [PubMed] [Google Scholar]
- Taschner P. E., Huls P. G., Pas E., Woldringh C. L. Division behavior and shape changes in isogenic ftsZ, ftsQ, ftsA, pbpB, and ftsE cell division mutants of Escherichia coli during temperature shift experiments. J Bacteriol. 1988 Apr;170(4):1533–1540. doi: 10.1128/jb.170.4.1533-1540.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Yousif S. Y., Broome-Smith J. K., Spratt B. G. Lysis of Escherichia coli by beta-lactam antibiotics: deletion analysis of the role of penicillin-binding proteins 1A and 1B. J Gen Microbiol. 1985 Oct;131(10):2839–2845. doi: 10.1099/00221287-131-10-2839. [DOI] [PubMed] [Google Scholar]
- Zijderveld C. A., Aarsman M. E., den Blaauwen T., Nanninga N. Penicillin-binding protein 1B of Escherichia coli exists in dimeric forms. J Bacteriol. 1991 Sep;173(18):5740–5746. doi: 10.1128/jb.173.18.5740-5746.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
- den Blaauwen T., Aarsman M., Nanninga N. Interaction of monoclonal antibodies with the enzymatic domains of penicillin-binding protein 1b of Escherichia coli. J Bacteriol. 1990 Jan;172(1):63–70. doi: 10.1128/jb.172.1.63-70.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
- van Heijenoort Y., Gómez M., Derrien M., Ayala J., van Heijenoort J. Membrane intermediates in the peptidoglycan metabolism of Escherichia coli: possible roles of PBP 1b and PBP 3. J Bacteriol. 1992 Jun;174(11):3549–3557. doi: 10.1128/jb.174.11.3549-3557.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]