Date of Award


Document Type

Open Access Dissertation

Degree Name

Doctor of Philosophy in Chemistry (PhD)

Administrative Home Department

Department of Chemistry

Advisor 1

Ashutosh Tiwari

Committee Member 1

Qinghui Chen

Committee Member 2

Haiying Liu

Committee Member 3

Lynn R. Mazzoleni


Proteins are one of the most versatile macromolecules in the biological system. The function or activity of a protein highly depends on its 3D native structure. However, under stress, they are at risk of misfolding/aggregation, leading to formation of structures that can indicate loss of function or gain of toxicity. In severe cases, protein aggregation can result in many diseases, including neurodegenerative diseases, such as Alzheimer’s, Parkinson’s, Huntington’s, and amyotrophic lateral sclerosis. Due to the heterogeneous nature of cellular environment and protein molecules, mechanism of in vivo folding and related toxicity still remains elusive. To have a better understanding of the cellular protein aggregations process and subsequent toxicity, we have performed aggregation studies of proteins with different types of posttranslational modifications, which is critical to protein’s functional diversity.

In this dissertation, two common types of covalent modification of proteins, i.e. disulfide reduction and acetylation, were selected. In aggregation studies of two globular proteins, hen egg white lysozyme and bovine serum albumin (BSA), formation of amorphous aggregates were observed as a consequence of disulfide bond scrambling. The structural properties of the observed aggregates were distinct and depended on disulfide reduction level. In study of amyloid β (Aβ) peptide, the major pathological protein in Alzheimer’s disease, effect of acetylation of the two lysine (K) positions, K16 and K28, on protein aggregation were investigated. We observed that acetylation on K16 can significantly increase hydrophobicity of Aβ and disrupt amyloid fibril formation. Interestingly, the heterogeneous mixtures of wild type and acetylated peptides displayed increased cytotoxicity compared to the homogeneous samples. To further understand the toxicity of protein aggregates, we then compared the cytotoxicity of eleven different aggregates from lysozyme and BSA, varying in morphology, size, flexibility, and hydrophobicity. The results suggest that the protein conformational changes in the early stage of aggregation process are essential for a gain in toxicity. Thy observed toxic species are structurally flexible, however, no clear correlation was found between cytotoxicity and hydrophobicity. Considering all the toxicity results of Aβ peptide, lysozyme, and BSA, we noticed that mixtures of native and modified proteins or aggregates are usually highly toxic. Therefore, the observed cytotoxicity of different structures may result from the heterogeneity of samples that are flexible rather than any defined structure. Further analysis of the toxic conformation would require high resolution structure determination of different aggregated protein species.