Author Correction: Proteome-wide structural changes measured with limited proteolysis-mass spectrometry: an advanced protocol for high-throughput applications

Inositol hexakisphosphate kinases (IP6Ks) are emerging as relevant pharmacological targets because a multitude of disease-related phenotypes has been associated with their function. While the development of potent IP6K inhibitors is gaining momentum, a pharmacological tool to distinguish the mammalian isozymes is still lacking. Here, we implemented an analog-sensitive approach for IP6Ks and performed a high-throughput screen to identify suitable lead compounds. The most promising hit, FMP 201300, exhibited high potency and selectivity towards the unique valine gatekeeper mutants of IP6K1 and IP6K2, compared to the respective wild-type kinases. Biochemical validation experiments revealed an allosteric mechanism of action that was corroborated by HDX-MS measurements. The latter analysis suggested that displacement of the αC helix, caused by the gatekeeper mutation, facilitates the binding of FMP-201300 to an allosteric pocket adjacent to the ATP binding site. FMP-201300 therefore serves as a valuable springboard for the further development of compounds that can selectively target the three mammalian IP6Ks; either as analog-sensitive kinase inhibitors or as an allosteric lead compound for the wild-type kinases.

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An unconventional gatekeeper mutation sensitizes inositol hexakisphosphate kinases to an allosteric inhibitor

Aguirre

Hostachy

Dornan

et al. 2023

Preprint

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On the pH-dependence of α-synuclein amyloid polymorphism and the role of secondary nucleation in seed-based amyloid propagation

Frey

Ghosh

Qureshi

et al. 2023

Preprint

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The aggregation of the protein alpha-synuclein is closely associated with several neurodegenerative disorders and as such the structures of the amyloid fibril aggregates have high scientific and medical significance. However, it seems that there are about as many unique atomic-resolution structures of these aggregates as there are publications describing them. Obviously, this highly polymorphic nature of alpha-synuclein fibrils hampers efforts in disease-relevant in vitro studies on alpha-synuclein amyloid aggregation. In order to better understand the factors that affect polymorph selection we studied the structures of alpha-synuclein fibrils in vitro as a function of pH and buffer using cryo-EM helical reconstruction. We find that in the physiological range of pH 5.8-7.4 a pH-dependent selection between Types 1, 2 and 3 polymorphs occurs. Our results indicate that even in the presence of seeds, the polymorph selection during aggregation is highly dependent on the buffer conditions, attributed to the polymorph-unspecific nature of secondary nucleation. We also uncovered two new polymorphs that occur at pH 7.0 in phosphate buffered saline. The first is a monomeric Type 1 which highly resembles the structure of the juvenile-onset synucleinopathy polymorph found in patient-derived material. The second is a new Type 5 polymorph that resembles a polymorph that has been recently reported in a study that used diseased tissues to seed aggregation. Taken together, our results highlight the shallow amyloid energy hypersurface that can be altered by subtle changes in the environment, including the pH which is shown to play a major role in polymorph selection and in many cases appears to be the determining factor in seeded aggregation. The results also confirm the possibility of producing disease relevant structure in vitro.

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Author Correction: Proteome-wide structural changes measured with limited proteolysis-mass spectrometry: an advanced protocol for high-throughput applications

Cited by 2 publications

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An unconventional gatekeeper mutation sensitizes inositol hexakisphosphate kinases to an allosteric inhibitor

An unconventional gatekeeper mutation sensitizes inositol hexakisphosphate kinases to an allosteric inhibitor

On the pH-dependence of α-synuclein amyloid polymorphism and the role of secondary nucleation in seed-based amyloid propagation

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