Biophysical Society Thematic Meeting - October 25-30, 2015

Polymers and Self Assembly: From Biology to Nanomaterials Poster Session II

13-POS Board 13 Synthesis of CCR6 Sulfopeptides to Probe Specific Contacts with the Chemokine CCL20 Marlon Dias 1 , Fabio C. Almeida 2 , Ana Paula Valente 2 , Viviane S. De Paula 1 . 2 Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil. 1 Universidade Federal do Rio de Janeiro-Polo Xerém, Rio de Janeiro, Brazil, Chemokines are small soluble proteins that stimulate chemotactic cell migration through interactions with chemokine receptors, a family of G protein-coupled receptors expressed in leukocyte cell membranes. In addition to their inherent pro-inflammatory activity, chemokines also contributes to many pathologies including auto-immune diseases and cancer. The N- terminus domains of the chemokine receptors contain tyrosine amino acid residues. Tyrosine sulfation is a post translational modification that enhances protein-protein interactions and the sulfation in the N-terminus regions of the chemokine receptors enhance the binding affinity of chemokine ligands and can modify the selectivity of a receptor. The aim of this work is characterize the interactions of the chemokine CCL20 with several CCR6-derived extracellular peptides by NMR spectroscopy. In this work we showed a strategy for synthesis of peptides containing sulfotyrosine at one or more specific positions to explore the role of tyrosine sulfation in recognition of the CCL20 by its receptor CCR6. We report the synthesis, purification and analyses of peptides corresponding to residues 16-30 of CCR6 in sulfated and non-sulfated form at the Y18, Y26 and Y27 positions. The non-sulfated peptide and sulfopeptide (sY27) were obtained with success and with high yield. The samples exhibited a major peak on preparative HPLC chromatograms and the correct masses of both peptides were verified by electrospray ionization mass spectrometry. The peptide resonances were assigned by analysis of TOCSY spectra. Next, we will demonstrate that sulfation of this peptide at one or both positions would substantially enhance the affinity of chemokine binding. Our objective is understand which sulfotyrosine present in the CCR6 sulfopeptide is critical for CCL20 recognition and highlights the potential influence of initial binding interactions on receptor activation.

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