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Structural analysis of SGNH domains involved in lipopolysaccharide modification

Tindall, Sarah (2017) Structural analysis of SGNH domains involved in lipopolysaccharide modification. MSc by research thesis, University of York.

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Gram negative bacteria have lipopolysaccharides (LPS) projecting from their outer membrane which are often acetylated by acyltransferase proteins. Some of these acyltransferase proteins have additional attached SGNH domains (SGNHAT3); while much is known about isolated SGNH domains (SGNHisol), little is known about those attached to acyltransferases. The aim of this research was to determine any structural or mechanistic differences between SGNHAT3 and SGNHisol domains. In silico analysis showed that while the catalytic residues are present in SGNHAT3 domains, the oxyanion hole residues, normally conserved in SGNHisol, are not present. The SGNHAT3 domains of two acyltransferases from Salmonella ser. Paratyphi A and Neisseria meningitidis were expressed and the SGNH domain from Salmonella ser. Paratyphi A purified. Mass spectrometry showed that two disulfide bonds were present and circular dichroism determined that reduction of these disulfide bonds had no effect on the thermal stability of the protein. The structure was determined using X-ray crystallography and, although the overall fold was the same, there were many structural differences between this SGNHAT3 domain and SGNHisol domains. The linker, previously thought to be flexible, was shown to be part of the SGNHAT3 domain structure. An additional helix was seen, not present in SGNHisol domains, and this could potentially be important for interaction with the acyltransferase domain. In addition, docking models highlighted residues potentially important for binding, however, further analysis will need to be carried out to investigate this.

Item Type: Thesis (MSc by research)
Academic Units: The University of York > Biology (York)
Depositing User: Miss Sarah Tindall
Date Deposited: 19 Feb 2018 14:21
Last Modified: 19 Feb 2018 14:21
URI: http://etheses.whiterose.ac.uk/id/eprint/19154

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