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Effect of macrocyclization and tetramethylrhodamine labeling on chemokine binding peptides

Wack, Julia S. ; Brahm, Kevin ; Babel, Philipp ; Dalton, James A. R. ; Schmitz, Katja (2023)
Effect of macrocyclization and tetramethylrhodamine labeling on chemokine binding peptides.
In: Journal of Peptide Science, 29 (7)
doi: 10.1002/psc.3486
Artikel, Bibliographie

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Kurzbeschreibung (Abstract)

Receptor‐derived peptides have played an important role in elucidating chemokine‐receptor interactions. For the inflammatory chemokine CXC‐class chemokine ligand 8 (CXCL8), a site II‐mimetic peptide has been derived from parts of extracellular loops 2 and 3 and adjacent transmembrane helices of its receptor CXC‐class chemokine receptor 1 (Helmer et al., RSC Adv., 2015, 5, 25657). The peptide sequence with a C‐terminal glutamine did not bind to CXCL8, whereas one with a C‐terminal glutamate did but with low micromolar affinity. We sought to improve the affinity and protease stability of the latter peptide through cyclization while also cyclizing the former for control purposes. To identify a cyclization strategy that permits a receptor‐like interaction, we conducted a molecular dynamics simulation of CXCL8 in complex with full‐length CXC‐class chemokine receptor 1. We introduced a linker to provide an appropriate spacing between the termini and used an on‐resin side‐chain‐to‐tail cyclization strategy. Upon chemokine binding, the fluorescence intensity of the tetramethylrhodamine (TAMRA)‐labeled cyclic peptides increased whereas the fluorescence anisotropy decreased. Additional molecular dynamics simulations indicated that the fluorophore interacts with the peptide macrocycle so that chemokine binding leads to its displacement and observed changes in fluorescence. Macrocyclization of both 18‐amino acid‐long peptides led to the same low micromolar affinity for CXCL8. Likewise, both TAMRA‐labeled linear peptides interacted with CXCL8 with similar affinities. Interestingly, the linear TAMRA‐labeled peptides were more resistant to tryptic digestion than the unlabeled counterparts, whereas the cyclized peptides were not degraded at all. We conclude that the TAMRA fluorophore tends to interact with peptides altering their protease stability and behavior in fluorescence‐based assays.

Typ des Eintrags: Artikel
Erschienen: 2023
Autor(en): Wack, Julia S. ; Brahm, Kevin ; Babel, Philipp ; Dalton, James A. R. ; Schmitz, Katja
Art des Eintrags: Bibliographie
Titel: Effect of macrocyclization and tetramethylrhodamine labeling on chemokine binding peptides
Sprache: Englisch
Publikationsjahr: 2023
Verlag: John Wiley & Sons
Titel der Zeitschrift, Zeitung oder Schriftenreihe: Journal of Peptide Science
Jahrgang/Volume einer Zeitschrift: 29
(Heft-)Nummer: 7
Kollation: 15 Seiten
DOI: 10.1002/psc.3486
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Kurzbeschreibung (Abstract):

Receptor‐derived peptides have played an important role in elucidating chemokine‐receptor interactions. For the inflammatory chemokine CXC‐class chemokine ligand 8 (CXCL8), a site II‐mimetic peptide has been derived from parts of extracellular loops 2 and 3 and adjacent transmembrane helices of its receptor CXC‐class chemokine receptor 1 (Helmer et al., RSC Adv., 2015, 5, 25657). The peptide sequence with a C‐terminal glutamine did not bind to CXCL8, whereas one with a C‐terminal glutamate did but with low micromolar affinity. We sought to improve the affinity and protease stability of the latter peptide through cyclization while also cyclizing the former for control purposes. To identify a cyclization strategy that permits a receptor‐like interaction, we conducted a molecular dynamics simulation of CXCL8 in complex with full‐length CXC‐class chemokine receptor 1. We introduced a linker to provide an appropriate spacing between the termini and used an on‐resin side‐chain‐to‐tail cyclization strategy. Upon chemokine binding, the fluorescence intensity of the tetramethylrhodamine (TAMRA)‐labeled cyclic peptides increased whereas the fluorescence anisotropy decreased. Additional molecular dynamics simulations indicated that the fluorophore interacts with the peptide macrocycle so that chemokine binding leads to its displacement and observed changes in fluorescence. Macrocyclization of both 18‐amino acid‐long peptides led to the same low micromolar affinity for CXCL8. Likewise, both TAMRA‐labeled linear peptides interacted with CXCL8 with similar affinities. Interestingly, the linear TAMRA‐labeled peptides were more resistant to tryptic digestion than the unlabeled counterparts, whereas the cyclized peptides were not degraded at all. We conclude that the TAMRA fluorophore tends to interact with peptides altering their protease stability and behavior in fluorescence‐based assays.

Freie Schlagworte: binding assay, chemokine‐receptor interaction, peptide macrocycles, stability, TAMRA labeling
ID-Nummer: e3486
Sachgruppe der Dewey Dezimalklassifikatin (DDC): 500 Naturwissenschaften und Mathematik > 540 Chemie
500 Naturwissenschaften und Mathematik > 570 Biowissenschaften, Biologie
Fachbereich(e)/-gebiet(e): 10 Fachbereich Biologie
10 Fachbereich Biologie > Computational Biology and Simulation
07 Fachbereich Chemie
07 Fachbereich Chemie > Clemens-Schöpf-Institut > Fachgebiet Biochemie
07 Fachbereich Chemie > Clemens-Schöpf-Institut > Fachgebiet Biochemie > Biologische Chemie
07 Fachbereich Chemie > Clemens-Schöpf-Institut
Hinterlegungsdatum: 31 Jul 2024 10:19
Letzte Änderung: 07 Aug 2024 09:56
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