A disintegrin derivative as a case study for PHIP labeling of disulfide bridged biomolecules

A specific labeling strategy for bioactive molecules is presented for eptifibatide (integrilin) an antiplatelet aggregation inhibitor, which derives from the disintegrin protein barbourin in the venom of certain rattlesnakes. By specifically labeling the disulfide bridge this molecule becomes access...

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Bibliographic Details
Published in:Scientific reports 2022-02, Vol.12 (1), p.2337-8, Article 2337
Main Authors: Fleckenstein, Max, Herr, Kevin, Theiß, Franziska, Knecht, Stephan, Wienands, Laura, Brodrecht, Martin, Reggelin, Michael, Buntkowsky, Gerd
Format: Article
Language:English
Subjects:
639/638/440/56
639/638/440/94
639/638/45/611
Disulfide bonds
High-performance liquid chromatography
Humanities and Social Sciences
Hyperpolarization
Labeling
Liquid chromatography
multidisciplinary
NMR
Nuclear magnetic resonance
Protein structure
Science
Science (multidisciplinary)
Tertiary structure
Venom
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Summary:A specific labeling strategy for bioactive molecules is presented for eptifibatide (integrilin) an antiplatelet aggregation inhibitor, which derives from the disintegrin protein barbourin in the venom of certain rattlesnakes. By specifically labeling the disulfide bridge this molecule becomes accessible for the nuclear spin hyperpolarization method of parahydrogen induced polarization (PHIP). The PHIP-label was synthesized and inserted into the disulfide bridge of eptifibatide via reduction of the peptide and insertion by a double Michael addition under physiological conditions. This procedure is universally applicable for disulfide-containing biomolecules and preserves their tertiary structure with a minimum of change. HPLC and MS spectra prove the successful insertion of the label. 1 H-PHIP-NMR experiments yield a factor of over 1000 as lower limit for the enhancement factor. These results demonstrate the high potential of the labeling strategy for the introduction of site selective PHIP-labels into biomolecules’ disulfide bonds.
ISSN:2045-2322
2045-2322
DOI:10.1038/s41598-022-06327-z