Investigating the role of helix-1 in the fibrillization of SAA1-76

PROJECT SUMMARY/ABSTRACT AA amyloidosis is a severe complication of chronic inflammatory disorders and is potentially fatal. The amyloid fibrils involved in AA amyloidosis are derived from serum amyloid A (SAA), which is an acute phase reactant protein. In AA amyloidosis, circulating amyloid fibrils are deposited in organs, ultimately leading to their

failure. Serum amyloid A (SAA) is the major protein component of amyloid fibrils associated with AA amyloidosis. There is accruing evidence that overexpression of SAA is correlated with the severity of inflammation caused by COVID-19. These highlight the need to unravel the mechanism of SAA misfolding that could in turn facilitate the

design of potential inhibitors. Molecular dynamic (MD) simulations have yielded an in silico model for SAA misfolding where the N-terminal helical (helix-1) region plays a pivotal role. Helix-1 is believed to transition from a -helix to -hairpin. Once formed, the -hairpin units are postulated to associate and stack upon themselves

to yield fibrils. We propose to investigate the veracity of the in silico model of SAA misfolding by synthesizing and characterizing peptide mutants derived from the helix-1 region of SAA and the longer fragment, SAA1-76. The roles of specific amino acids will be probed along with the trajectory each takes during SAA misfolding. Using

standard spectroscopic methods, we will determine changes in amyloidogenic propensity, conformation, key molecular interaction, and amyloid morphology of each peptide during or after SAA fibrillization. Experimental data will be correlated with the findings of the in silico results to yield information on the mechanism. The data

obtained from helix-1 fragments SAA1-13 and SAA1-27 will be extended to the truncated peptide, SAA1-76, found in amyloid deposits of SAA. We intend to emply in silico and in vitro study to help yield important mechanistic clues. One of the major goals of this project is to provide students, >50% of whom are underrepresented minorities,

with the opportunity to conduct research and gain experience working in a modern biomedical laboratory. Undergraduate researchers involved in the project will be expected to synthesize and spectroscopically characterize peptides, verify spectroscopic data using MD simulation, and characterize amyloid morphology.

Each student will focus on aspects of the project that best interest them and fit their background. Regular research group meetings will ensure that students participate in the planning of experiments and analysis of data. Students will have the opportunity to apply theories learned in the classroom to real world biochemical

investigation. Research experiences that are based on current technology allow students to solidify and fortify their knowledge. Hands-on experiential learning is one of the most effective approaches to develop students’ critical thinking, perseverance, and confidence that could translate to a fulfilling career. Their experience will help

them enter graduate programs and/or find employment in biomedical careers. The educational impact of this project also extends to local high school and community college students working in the PI’s laboratory. The research experiences of these students will help ensure the vibrancy of the scientific infrastructure in the US.