Peptides derived from CXCL8 based on in silico analysis inhibit CXCL8 interactions with its receptor CXCR1.

Shinn-Jong Jiang,Chun-Chun Chang,Je-Wen Liou,Hao-Jen Hsu,Yi Chung,Lee-Fong Lin

doi:10.1038/srep18638

Abstract

Chemokine CXCL8 is crucial for regulation of inflammatory and immune responses via activating its cognate receptor CXCR1. In this study, molecular docking and binding free energy calculations were combined to predict the initial binding event of CXCL8 to CXCR1 for peptide drug design. The simulations reveal that in the initial binding, the N-loop of CXCL8 interacts with the N-terminus of CXCR1, which is dominated by electrostatic interactions. The derived peptides from the binding region of CXCL8 are synthesized for further confirmation. Surface plasmon resonance analyses indicate that the CXCL8 derived peptide with 14 residues is able to bind to the receptor CXCR1 derived peptide with equilibrium KD of 252 μM while the peptide encompassing a CXCL8 K15A mutation hardly binds to CXCR1 derived peptide (KD = 1553 μM). The cell experiments show that the designed peptide inhibits CXCL8-induced and LPS-activated monocytes adhesion and transmigration. However, when the peptides were mutated on two lysine residues (K15 and K20), the inhibition effects were greatly reduced indicating these two amino acids are key residues for the initial binding of CXCL8 to CXCR1. This study demonstrates that in silico prediction based functional peptide design can be effective for developing anti-inflammation drugs.

Highlights

Excessive or prolonged leukocyte related inflammation generally leads to tissue destruction, which highlights the importance of properly controlling this inflammatory process
We recently proposed that CXCL8 binding to CXCR1 is a multistep process, which is in accordance with previous experiments[19]
Following the protocol of our pervious study[19], CXCR1 was constructed by combining the NMR experiment (PDB: 2LNL) and homology modeling of the N- and C-terminal parts

Summary

Introduction

Excessive or prolonged leukocyte related inflammation generally leads to tissue destruction, which highlights the importance of properly controlling this inflammatory process. Mutagenesis studies have demonstrated that charged residues near the third and fourth extracellular loops (EC loops) of CXCR1 are crucial for these interactions[11,12,13]. Based on these studies, a mechanism by which CXCL8 and CXCR1 interact has been proposed as occurring in a two-sites multistep process[12,14,15,16,17,18,19]. This study demonstrated an effective process for developing peptide drugs with inhibitory functions by using molecular docking predictions, binding free energy calculations, SPR measurements, and in vitro cellular assays

Methods

Results

Discussion

Conclusion