The study of two genes involved in the maintenance of intracellular redox potentials in Thiobacillus ferrooxidans
type: CreativeWork
description: The thioredoxin and ϒ-glutamylcysteine synthetase genes of Thiobacillus ferrooxidans were isolated by complementation of an Escherichia coli trxA gshA double mutant for growth on minimal medium lacking glutathione. Transduction of a T. ferrooxidans genome cos mid library into the E. coli mutant resulted in the identification of two groups of complementing cosmids. One of these groups was able to complement the TrxA⁺ requirement of an E. coli trxA met mutant for conversion of methionine sulfoxide to methionine. The nucleotide sequence of a 1.1 kbp Hindiii-Psti T.ferrooxidans chromosomal DNA fragment containing the cloned trxA gene, was determined. An open reading frame (ORF) corresponding to the thioredoxin gene and part of an ORF corresponding to the N-terminal region of the rho gene were identified. A 14 kDa protein corresponding to the T. ferrooxidan thioredoxin was synthesised in an E. coli-derived in vitro transcription/translation system. The predicted amino acid sequence of the T.ferrooxidans trxA gene showed 70.6% identity to that of E. coli. The cloned T. ferrooxidans trxA gene was able to support T7 phage replication in an E. coli trxA mutant to almost the same level as a TrxA⁺ wild type strain. However, it was not able to satisfy the thioredoxin requirement of M13 phage. Crude extracts of the E. coli trxA gshA mutant containing the cloned T. ferrooxidans trxA gene were able to reduce insulin far more rapidly those that did not contain the cloned gene, indicating that the T.ferrooxidans thioredoxin has disulfide reductase activity. DNA:RNA hybridisation analysis was carried out on transcripts prepared from the E. coli trxA gshA double mutant, the mutant containing the cloned trxA gene and T. ferrooxidans cells. A single transcript of about 0.5 kbp was obtained for RNA from T. ferrooxidans cells. Several transcripts were produced from the cloned T. ferrooxidans trxA gene in E. coli. One of these transcripts corresponded in size to the 0.5 kbp transcript prod…
abstract: The thioredoxin and ϒ-glutamylcysteine synthetase genes of Thiobacillus ferrooxidans were isolated by complementation of an Escherichia coli trxA gshA double mutant for growth on minimal medium lacking glutathione. Transduction of a T. ferrooxidans genome cos mid library into the E. coli mutant resulted in the identification of two groups of complementing cosmids. One of these groups was able to complement the TrxA⁺ requirement of an E. coli trxA met mutant for conversion of methionine sulfoxide to methionine. The nucleotide sequence of a 1.1 kbp Hindiii-Psti T.ferrooxidans chromosomal DNA fragment containing the cloned trxA gene, was determined. An open reading frame (ORF) corresponding to the thioredoxin gene and part of an ORF corresponding to the N-terminal region of the rho gene were identified. A 14 kDa protein corresponding to the T. ferrooxidan thioredoxin was synthesised in an E. coli-derived in vitro transcription/translation system. The predicted amino acid sequence of the T.ferrooxidans trxA gene showed 70.6% identity to that of E. coli. The cloned T. ferrooxidans trxA gene was able to support T7 phage replication in an E. coli trxA mutant to almost the same level as a TrxA⁺ wild type strain. However, it was not able to satisfy the thioredoxin requirement of M13 phage. Crude extracts of the E. coli trxA gshA mutant containing the cloned T. ferrooxidans trxA gene were able to reduce insulin far more rapidly those that did not contain the cloned gene, indicating that the T.ferrooxidans thioredoxin has disulfide reductase activity. DNA:RNA hybridisation analysis was carried out on transcripts prepared from the E. coli trxA gshA double mutant, the mutant containing the cloned trxA gene and T. ferrooxidans cells. A single transcript of about 0.5 kbp was obtained for RNA from T. ferrooxidans cells. Several transcripts were produced from the cloned T. ferrooxidans trxA gene in E. coli. One of these transcripts corresponded in size to the 0.5 kbp transcript prod…
author: Powles, Rosamund Eira
language: eng
date published: 1996
date modified: 2026-07-22T22:23:26Z
provider: University of Cape Town
publisher: Networked Digital Library of Theses and Dissertations
type: WebPage
source organization: Department of Molecular and Cell Biology

PropertyValue
property ID: Handle
value: http://hdl.handle.net/11427/17947