Identification of a novel carbohydrate-mimicking octapeptide from chemical peptide library and characterization as selectin inhibitor

Abstract

We found a novel octapeptide (H-YRNWFGRW-NH2) mimicking sialyl Lewis X (sLeX) carbohydrate from a chemical peptide library with anti-sLeX monoclonal antibody (MAb) 2H5. The peptide libraries were constructed by Fmoc-based solid-phase methodology using the mix-split method. The octapeptide sequence was determined by the iterative deconvolution method using anti-sLeX MAb 2H5. To define the important residues for interaction with anti-sLeX MAb 2H5, alanine-scanning analogues of H-YRNWFGRW-NH2 were synthesized. Substitution of Tyr1, Trp4, Arg7 and Trp8 to Ala resulted in a marked drop in affinity. This result indicates that aromatic and cationic amino residues have a key role in interacting with anti-sLeX MAb 2H5. The binding property of the octapeptide was evaluated with anti-sLeX MAb 2H5 and human E-selectin. The octapeptide showed high inhibitory potency (IC50=17.8 nM) for sLeX and competitively inhibited the binding of anti-sLeX MAb 2H5 in a dose-dependent manner. The octapeptide had high affinity (Kd=0.168 μM) for E-selectin and this binding was inhibited by sLeX. These results suggest that octapeptide binds to anti-sLeX MAb 2H5 or E-selectin at the sLe X binding site and sterically interferes with the recognition of anti-sLeX MAb 2H5 or E-selectin with sLeX. This peptide may be a useful lead compound for an anti-inflammatory agent targeting selectin. © 2011 Pharmaceutical Society of Japan.