Abstract
Legionella pneumophila infects eukaryotic cells by forming a replicative organelle – the Legionella containing vacuole. During this process, the bacterial protein DrrA/SidM is secreted and manipulates the activity and post-translational modification (PTM) states of the vesicular trafficking regulator Rab1. As a result, Rab1 is modified with an adenosine monophosphate (AMP), and this process is referred to as AMPylation. Here, we use a chemical approach to stabilise low-affinity Rab:DrrA complexes in a site-specific manner to gain insight into the molecular basis of the interaction between the Rab protein and the AMPylation domain of DrrA. The crystal structure of the Rab:DrrA complex reveals a previously unknown non-conventional Rab-binding site (NC-RBS). Biochemical characterisation demonstrates allosteric stimulation of the AMPylation activity of DrrA via Rab binding to the NC-RBS. We speculate that allosteric control of DrrA could in principle prevent random and potentially cytotoxic AMPylation in the host, thereby perhaps ensuring efficient infection by Legionella.
Highlights
Legionella pneumophila infects eukaryotic cells by forming a replicative organelle – the Legionella containing vacuole
Electrospray ionisation MS (ESI-MS) of the purified complex formed in living E. coli cells between R69BrC6KRab1b and D82CDrrA verified the identity of the thioether crosslinked complex, which is AMPylated at Y77Rab1b within Rab1b, confirming the enzyme and substrate activity of both DrrA and Rab1b mutants (Fig. 1d)
Infection of eukaryotic cells by L. pneumophila is a coordinated process characterised by the controlled release and activity regulation of bacterial proteins
Summary
Legionella pneumophila infects eukaryotic cells by forming a replicative organelle – the Legionella containing vacuole During this process, the bacterial protein DrrA/SidM is secreted and manipulates the activity and post-translational modification (PTM) states of the vesicular trafficking regulator Rab[1]. The formation and maintenance of the LCV are mediated by ~330 Legionella effector proteins that are released by the bacterial Type IVb secretion system (T4bSS) from the bacterium into the host. These effectors interfere with different host processes, including manipulation of vesicular trafficking The biology of Rab proteins has been the topic of a recent review[3]
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