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Title: Role of DiaA and SeqA homologues in the deep-sea adapted growth of Photobacterium profundum SS9
Author: El-Hajj, Ziad W.
ISNI:       0000 0004 2729 175X
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2009
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The mechanism of high pressure-adapted growth in the deep-sea bacterium Photobacterium profundum SS9 is poorly understood. To gain further insights, two P. profundum SS9R mutants were investigated. FL23 (pbpra3229::m-Tn10) and FL28 (pbpra1039::m-Tn10) had been previously characterised as high pressuresensitive and pressure-enhanced, respectively. FL23 had a growth defect at atmospheric pressure but failed to show high pressure-adapted growth on solid agar. Pbpra3229 is 75 % identical to E. coli DiaA (stimulator of DNA replication and critical for the timely initiation of replication) and 45% identical to E. coli GmhA (essential for lipopolysaccharide core biosynthesis), which led to an investigation into whether either process was affected in FL23. However, the lipopolysaccharide of FL23 and its parent strain were identical, which suggests that Pbpra3229 is not a GmhA homologue. In contrast, the pbpra3229 and E. coli diaA genes were functionally interchangeable and both restored the timing of DNA replication in an E. coli diaA mutant. FL28 had growth and morphological defects at high pressure, but both phenotypes were exacerbated at atmospheric pressure. Pbpra1039 is 55% identical to E. coli SeqA, which is a negative regulator of DNA replication and also essential for timely initiation. Pbpra1039 was shown to be a functional homologue of E. coli SeqA, as pbpra1039 partially complemented the DNA replication defect of an E. coli seqA mutant. Combined, these findings provide evidence that Pbpra3229 is a DiaA homologue, whereas Pbpra1039 is a cold adapted SeqA homologue, and that both positive and negative regulation of initiation of DNA replication are essential for the ability of P. profundum SS9 to adapt to deep-sea conditions. A marine metagenomic library was also screened for clones that produced novel cell envelope polysaccharides and tools were developed to identify cell envelope polysaccharides in P. profundum SS9.
Supervisor: Ferguson, Gail. ; Sawyer, Lindsay. Sponsor: Darwin Trust of Edinburgh ; Leverhulme Trust ; Royal Society ; Natural Environment Research Council (NERC)
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: photobacterium profundum ; high pressure-adapted growth