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Article Dans Une Revue Journal of Immunological Methods Année : 2002

Synthesis and characterization of biologically functional biotinylated RANTES

Résumé

Development of specifically labeled chemokines that retain their biological properties should be useful for analyzing their mechanisms of action both under physiological and pathological conditions. Here, we report the chemical synthesis and characterization of RANTES (regulated upon activation normal T cell expressed and secreted) derivatives that were biotinylated at residues 1, 25, 33, 45, or 67. Gel filtration and ultracentrifugation experiments showed that biotinylation at position 45 or 67 decreased the aggregation tendency of the chemokine to a dimeric state. Competition experiments, using a stably transfected CHO-K1 cell line overexpressing human CCR5, a RANTES receptor, indicated that derivatives biotinylated at positions 1, 25, and 67 bound to CCR5 with the same affinity as native RANTES. Flow cytometry analysis showed that RANTES biotinylated at residue 67 (B67-RANTES) bound more efficiently to primary macrophages than the other derivatives. Such binding was dependent on cell surface glycosaminoglycans (GAGs) since it was reduced when macrophages or HeLa cells expressing or not CCR5 were first treated with GAG-specific enzymes. In addition, B67-RANTES modulated CCR5 expression on lymphocytes and elicited chemotaxis of monocytes in the same manner as unmodified RANTES. Thus, B67-RANTES acts as a CCR5 agonist and may be useful to study the role of RANTES in pathologies such as, for example, human immunodeficiency virus (HIV) infection and inflammatory disorders.

Dates et versions

hal-02528550 , version 1 (01-04-2020)

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Claudio Vita, Eugenia Drakopoulou, Loyda Ylisastigui, Youssef Bakri, Jean Vizzavona, et al.. Synthesis and characterization of biologically functional biotinylated RANTES. Journal of Immunological Methods, 2002, 266 (1-2), pp.53-65. ⟨10.1016/s0022-1759(02)00096-0⟩. ⟨hal-02528550⟩
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