Synthesis of the chelator lipid nitrilotriacetic acid ditetradecylamine (NTA-DTDA) and its use with the IAsys biosensor to study receptor-ligand interactions on model membranes
Date
2001
Authors
Altin, Joseph
White, Felix
Easton, Christopher
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Publisher
Elsevier
Abstract
This work describes the synthesis and use of the chelator lipid, nitrilotriacetic acid ditetradecylamine (NTA-DTDA). This lipid is readily dispersed in aqueous media, both alone and when mixed with carrier lipids like dimyristoylphosphatidylcholine (DMPC). Fluorescence microscopic examination of membranes deposited from NTA-DTDA-containing liposomes shows that NTA-DTDA mixes uniformly with the carrier lipid, and does not phase separate. NTA-DTDA-membranes deposited onto the sensing surface of IAsys biosensor cuvettes show good stability, permitting use of the biosensor to study protein interactions. Hexahistidine-tagged proteins including recombinant forms of the extracellular regions of murine B7.1 (B7.1-6H) and of the human erythropoietin receptor (EPOR-6H) bind to NTA-DTDA-membranes; the stability of binding is dependent on both protein concentration, and density of NTA-DTDA. Kinetic measurements show that high stability of anchored proteins (t1/2 ∼ 10-20 h, apparent Kd ∼ 1 nM) can be achieved using membranes containing 25 mol% NTA-DTDA, but low levels of bound protein (< 200 arc seconds). The system is used to study the interaction of human EPO with the EPOR anchored onto NTA-DTDA-membranes. In addition to the biological applications reported recently, the results show that NTA-DTDA can be a useful reagent in the study of receptor-ligand interactions.
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Keywords
Keywords: acetic acid derivative; dimyristoylphosphatidylcholine; erythropoietin receptor; hexahistidine; histidine derivative; lipid; liposome; nitrilotriacetic acid ditetradecylamine; unclassified drug; article; artificial membrane; binding affinity; biosensor; c Chelator lipid; Erythropoietin; IAsys biosensor; Membrane; Nitrilotriacetic acid; Receptor-ligand interaction
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Source
Biochimica et Biophysica Acta: Biomembranes
Type
Journal article
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2037-12-31