Structural characterisation of Bdellovibrio bacteriovorus predatory peptidoglycan modifying proteins

Valdivia Delgado, M. S. ORCID: 0000-0003-0816-9209 (2023). Structural characterisation of Bdellovibrio bacteriovorus predatory peptidoglycan modifying proteins. University of Birmingham. Ph.D.

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Bdellovibrio bacteriovorus is an endobiotic bacterial predator that encodes multiple enzymes related to known peptidoglycan (PG) modifiers, which are necessary for predation to occur. Additionally, a plethora of cryptic proteins are expressed and upregulated during predation. It has been shown that L,D-transpeptidases (LDTs) are required for bdelloplast formation and stabilisation. These LDTs cluster in six groups in B. bacteriovorus HD100, nevertheless, no further LDT information has been previously described. Here we have determined and analysed the structures of LDTs Bd0566, Bd1075, Bd3334 and Bd3741, showing that even though they share a structurally conserved catalytic domain, they
have acquired accessory domains and motifs characteristic to each LDT group. Moreover, we have integrated these novel structures with the previously solved structures of Bd0553, Bd0886 and Bd1402, generating a panel of seven LDTs representing the six groups of B. bacteriovorus HD100, allowing us to propose possible roles and the structural determinants defining the transferase/crosslinking or hydrolytic activities of each group. Further, we report the structure of two predatory NlpC/P60 amidohydrolases/peptidases, Bd0601 and Bd1177. We have determined that Bd0601 is a DUF1460 amidohydrolase, wherein the PG stem peptide binding groove is blocked by a tyrosine residue, suggesting the need of a conformational rearrangement for substrate binding. On the other hand, we have determined that Bd1177 is a bilobular NlpC/P60, wherein the active site is located in the interface between the catalytic N-terminal domain and a novel C-terminal regulatory domain, suggesting that Bd1177 fluctuates to an open conformation upon substrate binding. Finally, we have determined the crystallographic structure of the cryptic secreted protein Bd0675, depicting a novel fold. The solved structure depicts a prominent central acidic and aromatic groove delimited by a conserved disulphide-stabilised loop, suggesting that this protein has ligand binding functions. Based on the solved structure we have tested different candidate ligands; however further experiments are required to propose a function for Bd0675. In addition, due to its novelty, Bd0675 was used as target for multiple structure predicting algorithms in CASP14.

Type of Work: Thesis (Doctorates > Ph.D.)
Award Type: Doctorates > Ph.D.
Licence: All rights reserved
College/Faculty: Colleges (2008 onwards) > College of Life & Environmental Sciences
School or Department: School of Bioscience
Funders: Other
Other Funders: ANID, Chile
Subjects: Q Science > Q Science (General)
Q Science > QH Natural history > QH301 Biology
Q Science > QR Microbiology


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