Sulfotyrosines are an integral part of biology; they are found in numerous systems and can augment intermolecular interactions. Chemokine receptors are a family of proteins that have sulfotyrosine modifications in their N-terminal region. Chemokine receptors are also good drug targets. Mutations that eliminated sulfotyrosines from the N-terminus of chemokine receptors have resulted in a range of negative biological responses such as reduction or loss of binding to its different ligands and reductions or loss of biological activities. So understanding the role of sulfotyrosine in chemokine receptors can improve drug design and expand the knowledge base. In vitro studies have shown that sulfotyrosine can enhance binding in two sulfopeptide systems. In this work, we show that sulfotyrosine can enhance binding in a third model system; between CCR3 N-terminal sulfopeptides and eotaxin-1. We also show that the amount of enhancement is dependent on the position of the sulfotyrosine. In addition, based on the dissociation constant, we found that in the absence of sulfotyrosine, CCR3 N-terminal peptide was unable to differentiate between eotaxin-1, -2, and -3. However, the addition of a single sulfotyrosine at position 16 allowed the CCR3 N-terminal peptide to bind to eotaxin-1 much more strongly than to eotaxin-2 or -3. Our results suggest that sulfotyrosine patterns may allow chemokine receptors to differentiate between their different ligands.