Substrate Recognition by the Legionella pneumophila Type II Secretion System.
Abstract
The bacterial Type II Secretion System (T2SS) is a transmembrane multiprotein assembly
that ejects folded protein substrates from the periplasm into the extracellular milieu. These
substrates are essential for pathogenesis in humans, animals, and plants. In the intracellular
pathogen Legionella pneumophila, the T2SS has been shown to play an important role in evading
recognition by the host during infection. Although much is known about the structures of the
compositional proteins of the system, recognition and recruitment of substrates for secretion by
the system is a poorly understood process. Previous studies have demonstrated binding between
the inner membrane protein XcpPGspC and T2SS substrates in Pseudomonas aeruginosa thus
implicating its involvement in this process. In this thesis, the periplasmic linker region of the
L. pneumophila homologue of XcpPGspC (LspCGspC) and the T2SS substrate Novel Type II
secreted protein A (NttA) (13 kDa) were recombinantly expressed and puri ed in Escherichia
coli. Using NMR spectroscopy titration experiments, a weak fast-exchange interaction between
these two proteins was observed, showing similar a nities to that previously demonstrated in
the study in P. aeruginosa. Solution structures of the two proteins were then solved by NMR
spectroscopy. Then, through the computational docking of these two proteins and guided by
NMR titration data, a recognition complex was characterised revealing residues essential in the
binding process. This is the rst discovered structure of a recognition complex between the
T2SS and its substrates in L. pneumophila. This study suggests a novel pharmacophore model
that may be used in antibiotic drug design of inhibitors to the formation of the recognition
complex.
Authors
Portlock, TheoCollections
- Theses [4235]