Biology subjects

Hamilton, A. D.

Publications and source records attributed to Hamilton, A. D..

3 recordsLinked to original sources

bioRxiv2022

Single-molecule nanopore dielectrophoretic trapping of α-Synuclein with lipid membranes

The lipid--Synuclein (-Syn) interaction plays a crucial role in the pathogenesis of Parkinsons disease. Here, we trap -Syn at a conjunction of an -hemolysin (HL) single nanopore-lipid to investigate the folding and unfolding kinetics of -Syn in a lipidic environment. The hybridized -Syn is generated through a reaction between a 5-thiol-modified nucleotide oligo (dC30) and the -Syn mutant (A140C). Owing to an applied voltage, single-molecule hybridized -Syn can be trapped at the single nanopore. The trapping events are associated with dielectrophoretic force. The folding and unfolding events of -Syn can be observed at the pore-membrane junction through interpretation of blockade current amplitudes and dwell time. This can be related to the protein quaternary structure influenced by the -Syn-membrane interaction, allowing further analysis of -Syn conformational dynamics. We studied how disease associated metal ions (Cu2+, Zn2+) modulate folding and unfolding of -Syn at the interface of the membranes and pore, and how -helical peptidomimetics stabilize the helical conformation of -Syn in the presence of a membrane. These studies aid our understanding of the complexity of the interaction of -Syn, lipid membranes and metal ions, and in using peptidomimetics, a new strategy against -Syn toxicity and aggregation is advanced.

biochemistry↗

bioRxiv2020

Protein mimetic amyloid inhibitor potently abrogates cancer-associated mutant p53 aggregation and restores tumor suppressor function

Missense mutations in p53 are severely deleterious and occur in over 50% of all human cancers. The vast majority of these mutations are located in the inherently unstable DNA-binding domain (DBD), many of which destabilize the domain further and expose its aggregation-prone hydrophobic core, prompting self-assembly of mutant p53 into inactive cytosolic amyloid-like aggregates. Screening an oligopyridylamide library, previously shown to inhibit amyloid formation associated with Alzheimers disease and type II diabetes, identified a tripyridylamide, ADH-6, that potently abrogates self-assembly of the aggregation-nucleating subdomain of mutant p53 DBD. Moreover, ADH-6 effectively targets and dissociates mutant p53 aggregates in human cancer cells, which restores p53s transcriptional activity, leading to cell cycle arrest and apoptosis. Notably, ADH-6 treatment substantially shrinks xenografts harboring mutant p53 and prolongs survival, while exhibiting no toxicity to healthy tissue. This study demonstrates the first successful application of a bona fide small-molecule amyloid inhibitor as an anticancer agent.

cancer biology↗

bioRxiv2017

Allosteric activation dictates PRC2 activity independent of its recruitment to chromatin

PRC2 is a therapeutic target for several types of cancers currently undergoing clinical trials. Its activity is regulated by a positive feedback loop whereby its terminal enzymatic product, H3K27me3, is specifically recognized and bound by an aromatic cage present in its EED subunit. The ensuing allosteric activation of the complex stimulates H3K27me3 deposition on chromatin. Here, we report a step-wise feedback mechanism entailing key residues within distinctive interfacing motifs of EZH2 or EED that are found mutated in cancers and/or Weaver syndrome. PRC2 harboring these EZH2 or EED mutants manifest little activity in vivo but, unexpectedly, exhibited similar chromatin association as wild-type PRC2, indicating an uncoupling of PRC2 activity and recruitment. With genetic and chemical tools, we further demonstrated that targeting allosteric activation overrode the gain-of-function effect of EZH2Y646X oncogenic mutations. These results revealed critical implications to the regulation and biology of PRC2 and a novel vulnerability in tackling PRC2-addicted cancers.

biochemistry↗