Disulfide bond formation plays a key role in the catalytic and regulatory functions of diverse proteins. Less known is its role in triggering aggregation and self-assembly of thiol-containing metabolites. These phenomena have been implicated in several metabolic disorders and neurodegenerative diseases such as kidney stone formation and more recently in the etiology of Alzheimer's disease (AD). In AD, an elevated level of homocysteine (Hcy), a metabolite of methionine, has been shown to promote the aggregation of AD-associated β-amyloid (Aβ) polypeptide. Hcy self-assembles into aggregates which interact with Aβ to alter its function and facilitate its deposition. Similarly, cysteine spontaneously forms cystine which further aggregates into the cystine kidney stones. Thus, it is important to study the aggregation of metabolites to capture the early onset of the diseases and develop effective therapy that targets metabolite aggregation.

In this project, we used ion mobility spectrometry-mass spectrometry (IMS-MS), powder X-ray diffraction, and biological assays to elucidate the self-assemblies of thiol-containing metabolites. Large oligomers were observed up to n = 36 for Hcy, all except the monomer were even-sized, implying disulfide bond formation stabilized oligomeric species. We observed that Hcy dimers (homocystine) assemble into tube-like structures consistent with powder X-ray diffraction data. Using transgenic mice, we were able to show that Hcy aggregates are co-localized with Aβ(1-42) fibrils.
In the case of cysteine/cystine, we probed further into their interactions with two commercial drugs, tioprinin and D-penicillamine, both were used in treatments of cystine kidney stones. We were able to show that tioprinin is the better drug. Finally, we showed that cystine aggregation can also be intervened by epigallocatechin gallate (a polyphenol) but via a different mechanism.

(A) Representative mass spectral data of Hcy showing various oligomers (i-v) (B) 2D plots of <i>m/z</i> vs. drift time (ms) for (i) <i>m/z</i> 939–943 and (ii) <i>m/z</i> 1345–1350 showing same isotope spacing in B(i) and overlapping isotope patterns in B(ii)

(A) Representative mass spectral data of Hcy showing various oligomers (i-v) (B) 2D plots of m/z vs. drift time (ms) for (i) m/z 939–943 and (ii) m/z 1345–1350 showing same isotope spacing in B(i) and overlapping isotope patterns in B(ii)

Thanh Do

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