Mass spectrometry analysis of human tissue to characterise cleavage events resulting in amyloid deposition associated with aortic aneurysm and dissection

The most common form of localised amyloid occurs in the aorta (aortic medial amyloid; AMA) and is estimated to occur in 97% of Caucasian people over 50. The main constituent of AMA has been identified to be fragments thought to be derived from the proteolysis of milk fat globule-EGF factor 8 protein (MFGE8). The full pathological impact of AMA is unknown, but it is believed that extracellular amyloid accumulation contributes to age-related diminished elasticity of the aorta and may underlie the pathogenesis of sporadic thoracic aortic aneurysm, in turn leading to rupture.

The aim of this project is to identify which MFGE8 fragments form amyloid deposits in human tissue obtained during aneurysm and dissection surgery at Liverpool Heart and Chest Hospital using mass spectrometry (MS)-based methods. Following this conceivable mechanisms of cleavage of MFGE8 that could result in fragments identified in human tissue will be probed with a view to manipulating cellular MFGE8 proteolysis and thereby reduce disease-causing AMA.

Amyloid will be isolated from human tissue using two approaches optimised in the host lab; extraction and laser microdissection. MS techniques, including top-down analysis of intact proteins and ion-mobility-MS, will be used to define the sequence of those protein fragments present within the amyloid deposits and explore aggregation mechanism of the MFGE8 fragments. Following characterisation of MFGE8 cleavage products, bioinformatic and biochemical strategies will be used to evaluate the potential of different proteases, to generate the active amyloid forming protein variants. Training will be provided in all biochemical, biophysical and MS techniques required for the project.

The student will also be integrated within the Liverpool Aorta Biomechanics and Biochemistry Research Group (http://labb-group.com/) and Centre for Proteome Research (https://www.liverpool.ac.uk/pfg/). The project is suited to a student with at least a good B.Sc. Upper Second in Biological or Life Sciences.