The characterisation of proteins key to the hunting and invasion of prey by Bdellovibrio bacteriovorus

Walters-Morgan, Hannah Louise ORCID: 0000-0001-9417-1000 (2022). The characterisation of proteins key to the hunting and invasion of prey by Bdellovibrio bacteriovorus. University of Birmingham. Ph.D.

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Abstract

The work presented in this thesis centres on proteins involved in the hunting and invasion of prey by Bdellovibrio bacteriovorus HD100 (Bdellovibrio), an Oligoflexia bacterium that predates Gram- negative bacteria (Hahn et al., 2017). In particular this work focuses on the characterisation of the Cache domain MCP family, Bd1081, Bd2503, Bd2504 and Bd3092 (as well as SKB1291214 homologue Bd1081\(^{S}\), and a family of 18 putative transcription factors (TFs) with a domain of unknown function (DUF4423) specifically, Bd1172, Bd1885, Bd2625 and Bd3720.

The primary aim of this thesis was the study of how Bdellovibrio uses chemotactic information to locate prey. Prior to this work Bdellovibrio was thought to locate prey cells using hydrodynamic forces, while chemotaxis was solely for the location of small essential molecules (Jashnsaz et al., 2017). However, the structural and biochemical characterisation of the Cache domain MCP family revealed them to be highly specific to only 1 or 2 ligands, where the ligands sensed by Bd2503 and Bd2504 are Gram-negative centric. This allowed us to develop two working hypotheses for the hunting of prey by Bdellovibrio, the ‘Chum in the Water’ and the ‘Lions at the Watering Hole’ Mechanisms, both of which are presented here.

The secondary focus of the work presented here was to characterise and determine the function of DUF4423. The 18 member TF family containing this domain within Bdellovibrio were originally thought to be involved in switching the gene expression from attack phase to growth phase upon prey invasion. By obtaining a partial structure of Bd2625 we were able to confirm DUF4423 contained a winged helix- turn-helix like motif within the domain, and combined with EMSAs this allowed us to attribute the function of DNA binding to the DUF4423 domain. EMSAs of Bd1172, Bd1885 and in particular Bd3720 demonstrated that this family did not bind DNA in a specific manner as initially hypothesised. This lead us to developing a new hypothesis for how these proteins function within Bdellovibrio; where they act as a set of novel nucleoid associated proteins that bind DNA in a non-sequence specific manner and regulate gene expression by chromatin remodelling.

Type of Work:

Thesis (Doctorates > Ph.D.)

Award Type:

Doctorates > Ph.D.

Supervisor(s):