5 The role of quorum-sensing in the virulence of Pseudomonas ...

has been proven experimentally to function (Redaschi, Bickle 1996). The frameshift
within E105leuX_mod occurs within the conserved protein domain as denoted by
the alignment between E105leuX_mod and EcoP15I mod, as it occurs at amino acid
437 which is within the first 484 amino acids. The residual 47 amino acids of the
conserved domain could be important for the protein to function as a methylase. The
residual amino acids outside of the mod conserved domain could also be essential
by adding stability for the secondary and tertiary structure of the protein. The amino
acids outside of the mod conserved domain could be also important because one of
the functions of a type III RM system mod protein is to bind to the res subunit to
enable the restriction activity of the type III RM system. This binding site is likely
to be outside of the mod conserved domain. This analysis suggests that
E105leuX_mod is non-functional.
The literature suggests that an RM system, with a functional res without a functional
mod would be lethal to a bacterium cell. This is because in absence of the
complementary mod, the res would restrict the bacterial DNA. In the current
situation, the type III RM system, if closely related to EcoP15I, has a number of fail
safes. Reaschi et al. (1996) reported that when the EcoP15I RM system is
introduced into a bacterium, the mod gene is expressed immediately and expression
of the res gene is suppressed until sufficient levels of mod is present within the
bacterium. The same research paper also specified that the res subunit did not
demonstrate any enzymatic activity unless it was complexed with the mod subunit,
adding a further level of protection. The mutation of the last 47 residual amino acids
within mod subunit, which are likely to be involved in the binding of res, suggests
there would be no functional restriction activity. The most important evidence is E.
coli E105 grows in culture despite being predicted to have a non-functional mod.
Further work to demonstrate that the mod gene was non-functional would be to
attempt to digest the genomic DNA from E105 with EcoP1I or EcoP15I but
unfortunately none was commercially available, neither was there a type II or type
III restriction enzyme with an identical recognition sequence commercially
available.
The predicted ORF E105leuX_1 translated to a protein that is 137 amino acids long.
This protein is predicted to have the putative conserved protein domain, COG3440
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