Journal article
Genetic Analysis of the Cell Division Protein FtsI (PBP3): Amino Acid Substitutions That Impair Septal Localization of FtsI and Recruitment of FtsN
Journal of bacteriology, Vol.186(2), pp.490-502
01/2004
DOI: 10.1128/JB.186.2.490-502.2004
PMCID: PMC305773
PMID: 14702319
Abstract
FtsI (also called PBP3) of
Escherichia coli
is a transpeptidase required for synthesis of peptidoglycan in the division septum and is one of several proteins that localize to the septal ring. FtsI comprises a small cytoplasmic domain, a transmembrane helix, a noncatalytic domain of unknown function, and a catalytic (transpeptidase) domain. The last two domains reside in the periplasm. We used PCR to randomly mutagenize
ftsI
, ligated the products into a green fluorescent protein fusion vector, and screened ∼7,500 transformants for
gfp-ftsI
alleles that failed to complement an
ftsI
null mutant. Western blotting and penicillin-binding assays were then used to weed out proteins that were unstable, failed to insert into the cytoplasmic membrane, or were defective in catalysis. The remaining candidates were tested for septal localization and ability to recruit another division protein, FtsN, to the septal ring. Mutant proteins severely defective in localization to the septal ring all had lesions in one of three amino acids—R23, L39, or Q46—that are in or near the transmembrane helix and implicate this region of FtsI in septal localization. Mutant FtsI proteins defective in recruitment of FtsN all had lesions in one of eight residues in the noncatalytic domain. The most interesting of these mutants had lesions at G57, S61, L62, or R210. Although separated by ∼150 residues in the primary sequence, these amino acids are close together in the folded protein and might constitute a site of FtsI-FtsN interaction.
Details
- Title: Subtitle
- Genetic Analysis of the Cell Division Protein FtsI (PBP3): Amino Acid Substitutions That Impair Septal Localization of FtsI and Recruitment of FtsN
- Creators
- Mark C Wissel - Department of Microbiology, University of Iowa, Iowa City, Iowa 52242David S Weiss - Department of Microbiology, University of Iowa, Iowa City, Iowa 52242
- Resource Type
- Journal article
- Publication Details
- Journal of bacteriology, Vol.186(2), pp.490-502
- DOI
- 10.1128/JB.186.2.490-502.2004
- PMID
- 14702319
- PMCID
- PMC305773
- NLM abbreviation
- J Bacteriol
- ISSN
- 0021-9193
- eISSN
- 1098-5530
- Publisher
- American Society for Microbiology
- Language
- English
- Date published
- 01/2004
- Academic Unit
- Microbiology and Immunology
- Record Identifier
- 9984001156702771
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