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Int. J. Mol. Sci. 2018, 19(3), 678; https://doi.org/10.3390/ijms19030678

Biochemical and MALDI-TOF Mass Spectrometric Characterization of a Novel Native and Recombinant Cystine Knot Miniprotein from Solanum tuberosum subsp. andigenum cv. Churqueña

1
Centro de Investigación de Proteínas Vegetales (CIPROVE), Departamento de Ciencias Biológicas, Facultad de Ciencias Exactas, Universidad Nacional de La Plata, 47 y 115 s/N, La Plata B1900AVW, Argentina
2
Institut de Biotecnologia i de Biomedicina, Universitat Autònoma de Barcelona, Campus Universitari, Bellaterra, Cerdanyola del Vallès, 08193 Barcelona, Spain
3
Catalan Institute of Nanoscience and Nanotechnology (ICN2), CSIC and The Barcelona Institute of Science and Technology, Campus UAB, Bellaterra, 08193 Barcelona, Spain
These authors contributed equally to this work.
*
Authors to whom correspondence should be addressed.
Received: 2 December 2017 / Revised: 11 January 2018 / Accepted: 16 January 2018 / Published: 28 February 2018
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Abstract

Cystine-knot miniproteins (CKMPs) are an intriguing group of cysteine-rich molecules that combine the characteristics of proteins and peptides. Typically, CKMPs are fewer than 50 residues in length and share a characteristic knotted scaffold characterized by the presence of three intramolecular disulfide bonds that form the singular knotted structure. The knot scaffold confers on these proteins remarkable chemical, thermal, and proteolytic stability. Recently, CKMPs have emerged as a novel class of natural molecules with interesting pharmacological properties. In the present work, a novel cystine-knot metallocarboxypeptidase inhibitor (chuPCI) was isolated from tubers of Solanum tuberosum, subsp. andigenum cv. Churqueña. Our results demonstrated that chuPCI is a member of the A/B-type family of metallocarboxypeptidases inhibitors. chuPCI was expressed and characterized by a combination of biochemical and mass spectrometric techniques. Direct comparison of the MALDI-TOF mass spectra for the native and recombinant molecules allowed us to confirm the presence of four different forms of chuPCI in the tubers. The majority of such forms have a molecular weight of 4309 Da and contain a cyclized Gln in the N-terminus. The other three forms are derived from N-terminal and/or C-terminal proteolytic cleavages. Taken together, our results contribute to increase the current repertoire of natural CKMPs. View Full-Text
Keywords: cystine-knot miniproteins; carboxypeptidase inhibitor; plant inhibitor; Solanum tuberosum; protease; Andean potatoes cystine-knot miniproteins; carboxypeptidase inhibitor; plant inhibitor; Solanum tuberosum; protease; Andean potatoes
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This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited (CC BY 4.0).

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Cotabarren, J.; Tellechea, M.E.; Tanco, S.M.; Lorenzo, J.; Garcia-Pardo, J.; Avilés, F.X.; Obregón, W.D. Biochemical and MALDI-TOF Mass Spectrometric Characterization of a Novel Native and Recombinant Cystine Knot Miniprotein from Solanum tuberosum subsp. andigenum cv. Churqueña. Int. J. Mol. Sci. 2018, 19, 678.

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