The Library
A study of the "ompT" gene of "Escherichia coli" K-12
Tools
Tools
Tools
Kemp, E. Helen (Elizabeth Helen) (1988) A study of the "ompT" gene of "Escherichia coli" K-12. PhD thesis, University of Warwick.
|
PDF
WRAP_Theses_KempH_1988.pdf - Submitted Version - Requires a PDF viewer. Download (5Mb) | Preview |
Official URL: http://webcat.warwick.ac.uk/record=b3229823~S15
Abstract
The ompT gene of E. coll encodes a 40 kd outer membrane protein (OmpT) which, in vitro, exhibits proteolytic activity towards the ferric-enterochelin receptor protein.
In this study the ompT gene was cloned from E. coli K-12 on a 4.3 kb EcoRI DNA-Fragment. Subcloning of this fragment, in conjunction with maxi-cell analysis, demonstrated that ompT was located on a 1.5 kb Pstl-Smal DNA fragment.
DNA sequence analysis revealed that the Pstl-Smal fragment contained an open reading frame (ORF) of 951 bp, the latter having a coding capacity of 35.6 kd. This deduced molecular weight was somewhat smaller than the molecular weight of 42 kd estimated by SDS-PAGE for pro-OmpT. Potential -10 and -35 promoter consensus sequences were identified upstream from the ompT coding region as was a putative ribosome binding site. A DNA sequence showing homology to a consensus sequence present in the putative promoter regions of iron- regulated genes, was also located upstream from the ompT coding region. With regard to the deduced amino acid sequence of OmpT, a potential signal sequence was found at the amino-terminal of the protein which could direct export from the cell cytoplasm. The protein was also examined for amino acid sequences displaying homology to other outer membrane proteins, but none were identified.
OmpT-phoA gene fusions were constructed in order to assess the ability of the ompT transcription/translation initiation signals to drive the expression of cloned heterologous genes in E. coli. OmpT-PhoA fusion proteins were indeed produced and these were exported to the periplasm of the cell. Synthesis of the chimeric proteins was found to be 5-6 fold higher at 37°C than at 30°C which seemed to reflect the normal temperature-dependent production of the OmpT protein. Both ompT-lacZ operon and protein fusions were also created in an attempt to define the stage at which temperature affected the synthesis of OmpT. Studies indicated that this occurred at some post-transcriptional step. The effect of the envZ allele on the expression of ompT was also investigated. This mutation appeared to reduce the level of transcription of ompT as Indicated by studies using the ompT-phoA and ompT-lacZ fusions.
| Item Type: | Thesis (PhD) | ||||
|---|---|---|---|---|---|
| Subjects: | Q Science > QR Microbiology | ||||
| Library of Congress Subject Headings (LCSH): | Escherichia coli, Escherichia coli -- Genetics | ||||
| Official Date: | 1988 | ||||
| Dates: |
|
||||
| Institution: | University of Warwick | ||||
| Theses Department: | Department of Biological Sciences | ||||
| Thesis Type: | PhD | ||||
| Publication Status: | Unpublished | ||||
| Supervisor(s)/Advisor: | Mann, Nicholas H. | ||||
| Sponsors: | Porton Down (Research Facility : Great Britain) | ||||
| Format of File: | |||||
| Extent: | xix, 268 leaves | ||||
| Language: | eng |
Request changes or add full text files to a record
Repository staff actions (login required)
 |
View Item |
Downloads
Downloads per month over past year
