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Site-directed mutagenesis. pcDNA3.1+ plasmids containing the human CCR2 mutants I208A5.45 and E291A7.39 (superscript indicates the Ballesteros Weinstein numbering system (Ballesteros, 1995), in which transmembrane residues are assigned two numbers that belong to the helix number and the residue number relative to the most conserved residue in this helix, which is assigned 50) with a 3x hemagglutinin (HA) epitope tag at the N-terminus were kindly provided by James Pease (Hall et al., 2009).
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