Beech, Matthew John (2021) Examining and exploiting docking domain-mediated carrier protein recognition in aeruginosin biosynthesis. PhD thesis, University of Warwick.

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Abstract

Nonribosomal-peptide synthetases (NRPSs) are multimodular megasynthetases that produce bioactive secondary metabolites in an assembly line-like manner. Selective protein-protein interactions between subunits in NRPS systems ensure biosynthetic fidelity and are typically mediated by recognition between C- and N-terminal docking domains. One example is that between beta-hairpin docking domains (βHDDs) and glutamate-rich short linear motifs (SLiMs).

Recent bioinformatics analysis has uncovered a novel βHDD-SLiM interface in NRPS subunits responsible for the biosynthesis of aeruginosin in Microcystis aeruginosa NIES-98 (scheme 1), which produces a mixture of non-, mono- and di-chlorinated aeruginosin congeners. In vitro reconstitution of the system has shown the AerA carrier protein domain bearing a C-terminal SLiM can interact productively with two distinct catalytic domains bearing Nterminal βHDDs, appended to the AerB condensation domain and a flavindependent halogenase, AerJ. Additionally, it has been demonstrated than competition between the two βHDDs for binding to the carrier protein SLiM is responsible for the mixture of chlorination patterns observed in the natural product.

Protein-protein interactions have been shown to be a key factor directing halogenase activity in the aeruginosin biosynthetic pathway. βHDD-bearing enzymes from evolutionarily divergent species can crosstalk productively with AerA(PCP) to generate novel molecules with desirable structural complexity. This work provides a new platform for pathway engineering by harnessing the potential of protein-protein interactions for the recruitment of carrier proteintethered substrates to direct natural product biosynthesis.

Item Type:	Thesis or Dissertation (PhD)
Subjects:	Q Science > QC Physics Q Science > QP Physiology
Library of Congress Subject Headings (LCSH):	Biosynthesis -- Research, Protein-protein interactions, Peptides, Ligases
Official Date:	March 2021
Dates:	Date Event March 2021 UNSPECIFIED
Institution:	University of Warwick
Theses Department:	Department of Chemistry
Thesis Type:	PhD
Publication Status:	Unpublished
Supervisor(s)/Advisor:	Challis, Gregory L. ; Lewandowski, Józef R.
Sponsors:	Midlands Integrative Biosciences Training Partnership ; Biotechnology and Biological Sciences Research Council (Great Britain)
Extent:	xxxii, 279 leaves : illustrations, charts
Language:	eng

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