The Glycobiology and Natural Products Chemistry of Prymnesium parvum

Abstract

The haptophyte microalga Prymnesium parvum is a globally important species due to the harmful algal blooms it forms, which have a devastating effect on the surrounding environment. A group of polyketide natural products, termed Prymnesins, are implicated in toxicity. Yet, despite decades of research, questions remain as to the abiotic and biotic factors modulating the toxicity of P. parvum. The role of nitrogen (N) and phosphorus (P) limitation, which has long been implicated in increased P. parvum toxicity, is first investigated. The increased abundance of Prymnesin-A1 under N or P limitation was confirmed, and other metabolome changes were examined. A bioinformatic based survey identified P. parvum as having an extensive capacity for natural product synthesis. Using two different transcriptomes, 8 non-ribosomal peptide synthetases (NRPSs) and 32 polyketide synthases (PKSs) were identified. Further investigation of one of the PKSs revealed a sequence of non-canonical domains likely involved in sugar nucleotide biosynthesis, which were further investigated. Intermediate capture probes, designed to investigate polyketide synthesis, have been successfully used In vivo with bacteria and fungi. Their suitability with microalgae was tested  with P. parvum, and Euglena gracilis. For P. parvum cultures the probe was highly toxic; E. gracilis cultures, on the other hand, survived probe incubation at all concentrations tested.   Lipidomics analysis of the cultures suggests the probes have an off-target effect in P. parvum. The sialic acid, 3-deoxy-D-glycero-D-galacto-2-nonulosonic acid (Kdn) is involved in viral infection in the haptophyte Emiliania huxleyi. The crystal structure of P. parvum CMP-Kdn synthetase is reported, and the role of a key active site residue in determining substrate specificity is experimentally validated. Phylogenetic analysis of CMP-sialic acid synthetases in microalgae concludes synthesis of Kdn is widespread. Given the presence of Kdn in P. parvum, the genome of its infecting virus, Prymnesium parvum DNA virus (PpDNAV) was surveyed for sugar-binding lectins. Four lectins were identified, and structural modelling with AlphaFold provided insight into their potential ligands. One lectin sequence was successfully expressed and its binding partner determined, indicating the likely role of lectins and sugars in algal-virus infection.

Date of Award	1 Aug 2024
Original language	English
Awarding Institution	The University of Manchester
Supervisor	Jason Micklefield (Supervisor) & Robert Field (Supervisor)