Domain-swapping analysis of FtsI, FtsL, and FtsQ, bitopic membrane proteins essential for cell division in Escherichia coli

Guzman, L. M.; Weiss, David S.; Beckwith, Jon

doi:10.1128/jb.179.16.5094-5103.1997

Cited by 95 publications

(131 citation statements)

References 42 publications

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“…Plasmids included pZAQ and pAQ (Yi et al, 1985) provided by J. Lutkenhaus, pBAD18Cm (Guzman et al, 1995) and pLMG161 (pBAD18ftsQ ) (Guzman et al, 1997) provided by L. M. Guzman and pGC401 . Plasmid pBAD18CmftsA was constructed by amplifying the ftsA gene from chromosomal DNA by PCR, surrounded by restriction sites for NheI (5Ј) and SphI (3Ј); the primers used were 5Ј-GGGGCTAGCGGAGCGGAGTAG-GCTGGGCG and 5Ј-GGGGCATGCACCTTCAATGCGCTCG-CGCA; the amplified fragment was cloned in NheI-and SphIdigested pBAD18Cm.…”

Section: Methodsmentioning

confidence: 99%

“…The resulting plasmid, pBAD18CmftsA, restores temperature resistance to an ftsA16 mutant. Plasmid pBADCmftsQ was constructed from pLMG161 (Guzman et al, 1997) by cloning the cat cassette of plasmid pACYC184, on a Bsa AI fragment, into the FspI site within the bla gene of pLMG161. The resulting plasmid, pBADCmftsQ, confers a Cm R Amp S phenotype (an active bla gene product confers mecillinam resistance).…”

Section: Methodsmentioning

confidence: 99%

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Analysis of the effect of ppGpp on the ftsQAZ operon in Escherichia coli

Navarro

Robin

D’Ari

et al. 1998

Molecular Microbiology

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Analysis of the effect of ppGpp on the ftsQAZ operon in Escherichia coli

Navarro

Robin

D’Ari

et al. 1998

Molecular Microbiology

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“…The resultant PCR product was digested with DpnI and was transformed into XL1-Blue. The plasmids pR 21 ⌽Lyz, in which the sequence encoding the N-terminal hydrophobic domain of the phage 21 endolysin R 21 replaced that of Lyz, pLyz⌽19, in which sequence encoding the N-terminal hydrophobic domain of Lyz was inserted between the first two codons of gene 19 from phage P22; and pLyz⌽PhoA, in which the signal sequence of PhoA was replaced with that of the N-terminal hydrophobic domain of Lyz, were constructed in a similar way. A cmyc-tagged allele of the R 21 gene was generated by using primers encoding the epitope flanked either with 15 nucleotides of homology 5Ј to the insertion site in R 21 or 15 nucleotides 3Ј to the insertion site.…”

Section: Methodsmentioning

confidence: 99%

“…The various endolysins, endolysin chimeras, and genes encoding FtsI and PhoA were placed under the control of the lac promoter of pJF118 (16). The DNA inserts for these constructs were PCR-amplified from the following sources: for pJFLyz, the P1 gene lyz was from P1vir DNA; for pJFR, the R gene was from pS105 (13); for pJF19, the P22 gene 19 was from P22 DNA; for pJFR 21 , the bacteriophage 21 R gene was from pBR121 (17); and for pJFFtsI and pJFPhoA, the ftsI and phoA genes were from E. coli chromosomal DNA. To construct pZAdsbA, the dsbA gene was PCR-amplified from E. coli chromosomal DNA.…”

Section: Methodsmentioning

confidence: 99%

A signal-arrest-release sequence mediates export and control of the phage P1 endolysin

Struck

Deaton

et al. 2004

Proc. Natl. Acad. Sci. U.S.A.

150

207

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The Lyz endolysin of bacteriophage P1 was found to cause lysis of the host without a holin. Induction of a plasmid-cloned lyz resulted in lysis, and the lytic event could be triggered prematurely by treatments that dissipate the proton-motive force. Instead of requiring a holin, export was mediated by an N-terminal transmembrane domain (TMD) and required host sec function. Exported Lyz of identical SDS͞PAGE mobility was found in both the membrane and periplasmic compartments, indicating that periplasmic Lyz was not generated by the proteolytic cleavage of the membrane-associated form. In gene fusion experiments, the Lyz TMD directed PhoA to both the membrane and periplasmic compartments, whereas the TMD of the integral membrane protein FtsI restricts Lyz to the membrane. Thus, the N-terminal domain of Lyz is both necessary and sufficient not only for export of this endolysin to the membrane but also for its release into the periplasm. The unusual N-terminal domain, rich in residues that are weakly hydrophobic, thus functions as a signal-arrest-release sequence, which first acts as a normal signal-arrest domain to direct the endolysin to the periplasm in membrane-tethered form and then allows it to be released as a soluble active enzyme in the periplasm. Examination of the protein sequences of related bacteriophage endolysins suggests that the presence of an N-terminal signalarrest-release sequence is not unique to Lyz. These observations are discussed in relation to the role of holins in the control of host lysis by bacteriophage encoding a secretory endolysin.

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“…Known fts mutants include ftsA, ftsI, ftsK, ftsL, ftsN, ftsW and ftsZ. [1][2][3][4][5][6][7][8] Inhibitors of protein synthesis exhibit bacteriostatic action except for the aminoglycoside (AG) antibiotics, 9) suggesting that the inhibition of protein synthesis by AG antibiotics is not directly involved in their bactericidal action. The bactericidal action of AG antibiotic is assumed to be based on their inhibition of DNA replication.…”

mentioning

confidence: 99%

Inhibition of Septation in Bacillus subtilis by a Peptide Antibiotic, Edeine B1

Shimotohno

Kawamura

Natori

et al. 2010

Biological & Pharmaceutical Bulletin

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The antibiotics, edeine A 1 and B 1 , are well-known inhibitors of protein biosynthesis in both prokaryote and eukaryote cells. Edeine A 1 and B 1 show almost the same biological activity. We show herein that edeine B 1 exhibits a lethal action in Bacillus subtilis, inhibiting septation via a mechanism different from the inhibition of protein synthesis by this antibiotic.The complex process of cell division is closely interconnected with other cellular processes. Chromosomal DNA replication, nuclear division, and cell division must be coordinated in time and space. DNA metabolism plays a pivotal role in the cell division events controlled by the partitioning of newly replicated chromosomes to daughter cells. This involves chromosomal DNA replication, the positioning of a septum at the midpoint of the cell, the assembly of cytoskeletal elements, and the cleavage of the septum after chromosome segregation. Genetic studies have unraveled the process of cell division by the analysis of filamenting temperaturesensitive mutants. In prokaryotes, filamenting temperaturesensitive (fts) genes produce the proteins essential for cell division. Known fts mutants include ftsA, ftsI, ftsK, ftsL, ftsN, ftsW and ftsZ. [1][2][3][4][5][6][7][8] Inhibitors of protein synthesis exhibit bacteriostatic action except for the aminoglycoside (AG) antibiotics, 9) suggesting that the inhibition of protein synthesis by AG antibiotics is not directly involved in their bactericidal action. The bactericidal action of AG antibiotic is assumed to be based on their inhibition of DNA replication. AG antibiotic blocks the initiation of DNA replication, 10) and interrupts oriC-membrane attachment. 11) The oriC-membrane attachment [12][13][14] is crucial for subsequent initiation of DNA replication, chromosome segregation, and septation of bacterial cells. Blocking DNA replication is known to lead to the save our ships (SOS) response in Escherichia coli, thereby inhibiting cell division. 15) We demonstrate herein that edeine B 1 inhibits cell division, exerting its bactericidal action by inhibiting FtsZ assembly, independently of its inhibitory effect on protein synthesis. MATERIALS AND METHODSPurification of Edeine B 1 Edeine B 1 was isolated from a culture filtrate of Bacillus brevis TTO2-8. The purified edeine B 1 yielded a single TLC spot, a single HPLC peak and the predicted structure based on NMR and FAB-MAS analysis. Edeine B 1 is a linear basic oligopeptide antibiotic consisting of five amino acid residues (isotyrosine, isoserine, a,b-diaminopropionic acid, 2,6-diamino-7-hydroxyazelanic acid, and glycine) and an organic base (guanylspermidine) produced by B. brevis. 16,17) Bacterial Strains B. subtilis 168 and mutant FtsZ ts1 (a temperature sensitive mutant of B. subtilis 168) were used.Viable Cell Number The viable cell number was counted by plating on LB (Luria-Bertani) agar, and recording the number of colonies that developed after 24 h incubation.Western Blotting Anti-FtsZ rabbit antibody raised against a chemically synthesized carboxyl t...

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Domain-swapping analysis of FtsI, FtsL, and FtsQ, bitopic membrane proteins essential for cell division in Escherichia coli

Cited by 95 publications

References 42 publications

Analysis of the effect of ppGpp on the ftsQAZ operon in Escherichia coli

Analysis of the effect of ppGpp on the ftsQAZ operon in Escherichia coli

A signal-arrest-release sequence mediates export and control of the phage P1 endolysin

Inhibition of Septation in Bacillus subtilis by a Peptide Antibiotic, Edeine B1

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