Exploring enzyme-mediated degradation of plant biomass by in situ imaging for biosecurity

It is estimated by the food and agriculture organisation that the global loss of crops is between 20-40%, costing over $220 billion in losses. These numbers will be exacerbated by climate change, which due to increased temperatures will drive pest proliferation. With the population expected to rise to 9.7 million by 2050, agriculture resilience needs to be understood on an atomic and cellular level.

Plant cell walls are composed of macro- and microfibrils of lignocellulose. Lignocellulose is comprised of a central rod of crystalline cellulose (β-1,4-linked glucose) surrounded by varying ratios of hemicellulose (polysaccharides of pyranoses and furanoses) and lignin (composed of phenolic compounds). The hydrogen bonding network of crystalline cellulose contributes to its recalcitrance. Glucose is used by many living organisms as an energy source, and to access glucose from cellulose, chewing and sucking herbivores, oomycete pathogens and fungal pathogens utilise lytic polysaccharide monooxygenases (LPMOs). LPMOs are a family of copper-dependent redox enzymes that break the β-1-4-glycosidic bonds between glucose molecules of cellulose using oxygen/hydrogen peroxide. They are categorised into 8 families based on their sequence within the CAZy database. Recently, these enzymes have been shown to be prominent virulent factors in pathogenic bacteria, fungi and oomycetes. Of the worldwide fungal pathogens and insect pests capable of infecting crop plants, multiple LPMO encoding genes have been identified.

The project will involve producing a variety of LPMO enzymes from bacterial, fungal and oomycetes pathogens and using matrix-assisted laser desorption ionisation (MALDI) and desorption electrospray ionisation (DESI) mass spectrometry imaging (MSI) to visualise, at the cellular level, the breakdown of plant cell wall matter. Additionally, laser ablation-inductively coupled plasma mass spectrometry imaging will be used to analyse the copper binding sites of LMPOs in pest-infected plant samples. Lastly, inhibition of these enzymes will be explored using the techniques mentioned.