The molecular basis for magnesium transport during virulence

Background - Pathogenicity islands containing genes associated with bacterial magnesium homeostasis have been shown to play a crucial role during virulence by Salmonella. Upon infection, the Salmonella bacteria initially encounters an acidic environment with low levels of Mg²⁺, conditions that lead to transcription of the genes,  mgtA and mgtB (Smith RL et al., 1998 Microbiolog). MgtA and MgtB belong to the P-type ATPases family of membrane transporters,  but are not essential for initial invasion, MgtB, does enhance the long term survival of the pathogen (Moncrief MB et al.,1999 J Biol Inorg Chem) while MgtA have been shown to enhance survival under high temperatures (O'Connor  K et al., 2009 PNAS). In addition, the virulence factor mgtC, which is essential for long-term survival in the host cell, is co-expressed with mgtB (Blanc-Potard AB  et la ., 1997 EMBO J).  It is established that MgtC does not transport Mg²⁺, but is a key player in inner macrophage survival (Alix  E et al., 2007 Trends Microbiol). MgtC is trafficked the inner membrane and believed to interact with the MgtB transport protein, although there is no experimental evidence for this direct protein-protein interaction. Interestingly there is also experimental findings suggesting that MgtC can modulate the activity of eukaryotic Na⁺, K⁺ -ATPase (Gunzel D et al., 2006 J Bacteriol). The purpose of this project is from a regulatory and structural point of view to investigate the molecular mechanism of Mg²⁺ transport and homeostasis in pathogenic bacteria, with the main focus on the system the P-type ATPases MgtA and MgtB is a part of.

Projects - We have expressed MgtA and established an activity assay to measure the reaction rate. Purification and Activity assay protocol for MgtB and MgtC, which are the genes located in the same operon

Molecular biologcal technicues you are likely to come into contact with and could be trained in

• Primer design, cloning, subcloning into E. coli.
• Expression of membrane proteins in E. coli.
• Purification and relipidering of membrane proteins.
• Characterization of membrane proteins such as phosphorylation experiments
• Crystallization and structure solution of proteins.
 

Figure 1, Functional characterization – Substrate specificity of MgtA

We are located in the Science Park on the 5th Floor in completely renovated laboratories. The general language in the laboratory is english

http://www.ncmm.uio.no/research/groups/membrane-transport/