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Open AccessArticle

Structural and Aggregation Features of a Human κ-Casein Fragment with Antitumor and Cell-Penetrating Properties

1
Institute of Chemical Biology and Fundamental Medicine SB RAS, Novosibirsk 630090, Russia
2
N.N. Vorozhtsov Novosibirsk Institute of Organic Chemistry SB RAS, Novosibirsk 630090, Russia
3
Department of Natural Sciences, Novosibirsk State University, 1 Pirogova Str., Novosibirsk 630090, Russia
*
Author to whom correspondence should be addressed.
Academic Editor: Hidekazu Hiroaki
Molecules 2019, 24(16), 2919; https://doi.org/10.3390/molecules24162919
Received: 29 June 2019 / Revised: 6 August 2019 / Accepted: 9 August 2019 / Published: 12 August 2019
(This article belongs to the Special Issue Protein Domains: Structures and Molecular Functions)
Intrinsically disordered proteins play a central role in dynamic regulatory and assembly processes in the cell. Recently, a human κ-casein proteolytic fragment called lactaptin (8.6 kDa) was found to induce apoptosis of human breast adenocarcinoma MCF-7 and MDA-MB-231 cells with no cytotoxic activity toward normal cells. Earlier, we had designed some recombinant analogs of lactaptin and compared their biological activity. Among these analogs, RL2 has the highest antitumor activity, but the amino acid residues and secondary structures that are responsible for RL2′s activity remain unclear. To elucidate the structure–activity relations of RL2, we studied the structural and aggregation features of this fairly large intrinsically disordered fragment of human milk κ-casein by a combination of physicochemical methods: NMR, paramagnetic relaxation enhancement (PRE), Electron Paramagnetic Resonance (EPR), circular dichroism, dynamic light scattering, atomic force microscopy, and a cytotoxic activity assay. It was found that in solution, RL2 exists as stand-alone monomeric particles and large aggregates. Whereas the disulfide-bonded homodimer turned out to be more prone to assembly into large aggregates, the monomer predominantly forms single particles. NMR relaxation analysis of spin-labeled RL2 showed that the RL2 N-terminal region, which is essential not only for multimerization of the peptide but also for its proapoptotic action on cancer cells, is more ordered than its C-terminal counterpart and contains a site with a propensity for α-helical secondary structure. View Full-Text
Keywords: intrinsically disordered protein; dimerization; casein micelle; disulfide bond; β-mercaptoethanol adduct intrinsically disordered protein; dimerization; casein micelle; disulfide bond; β-mercaptoethanol adduct
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Chinak, O.A.; Shernyukov, A.V.; Ovcherenko, S.S.; Sviridov, E.A.; Golyshev, V.M.; Fomin, A.S.; Pyshnaya, I.A.; Kuligina, E.V.; Richter, V.A.; Bagryanskaya, E.G. Structural and Aggregation Features of a Human κ-Casein Fragment with Antitumor and Cell-Penetrating Properties. Molecules 2019, 24, 2919.

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