Genetic studies on the region downstream of the unc operon of Thiobacillus ferroxidans

Master Thesis


Permanent link to this Item
Journal Title
Link to Journal
Journal ISSN
Volume Title

University of Cape Town

A Tn7-like element was found in a region downstream of a cosmid (p818.1) isolated from a genomic library of Thiobacillus ferrooxidans ATCC 33020. A probe made from the Tn7-like element hybridized to restriction fragments of identical size from both cosmid p818.1 and T ferrooxidans chromosomal DNA. The same probe hybridized to restricted chromosomal DNA from two other T ferrooxidans strains (ATCC 23270 and 19859). There were no positive signals when an attempt was made to hybridize the probe to chromosomal DNA from two Thiobacillus thiooxidans strains (ATCC 19733 and DSM504) and a Leptospirillum ferrooxidans strain DSM 2705. A 3.5 kb BamHI-BamHI fragment was subcloned from p818.1 downstream the T ferrooxidans unc operon and sequenced in both directions. One partial open reading frame (ORFl) and two complete open reading frames (ORF2 and ORF3) were found. On the basis of high homology to previously published sequences, ORFI was found to be the C-terminus of the T ferrooxidans glmU gene encoding the enzyme GlcNAc I-P uridyltransferase (EC The ORF2 was identified as the T ferrooxidans glmS gene encoding the amidotransferase, glucosamine synthetase (EC The third open reading frame (ORF3) was found to have very good amino acid sequence homology to TnsA of transposon Tn7. Inverted repeats very similar to the imperfect inverted repeat sequences of Tn7 were found upstream of ORF3. The cloned T ferrooxidans glmS gene was successfully used to complement an E.coli glmS mutant CGSC 5392 when placed behind a vector promoter, but was otherwise not expressed in E.coli.

Bibliography: pages 144-161.