N-Methyl mesoporphyrin IX (NMM) is exceptionally selective for G-quadruplexes (GQ) relative to duplex DNA and, as such, has found a wide range of applications in biology and chemistry. In addition, NMM is selective for parallel versus antiparallel GQ folds, as was recently demonstrated in our laboratory. Here, we present the X-ray crystal structure of a complex between NMM and human telomeric DNA dAGGG(TTAGGG)₃, Tel22, determined in two space groups, P2₁2₁2 and P6, at 1.65 and 2.15 Å resolution, respectively. The former is the highest resolution structure of the human telomeric GQ DNA reported to date. The biological unit contains a Tel22 dimer of 5′-5′ stacked parallel-stranded quadruplexes capped on both ends with NMM, supporting the spectroscopically determined 1:1 stoichiometry. NMM is capable of adjusting its macrocycle geometry to closely match that of the terminal G-tetrad required for efficient π–π stacking. The out-of-plane N-methyl group of NMM fits perfectly into the center of the parallel GQ core where it aligns with potassium ions. In contrast, the interaction of the N-methyl group with duplex DNA or antiparallel GQ would lead to steric clashes that prevent NMM from binding to these structures, thus explaining its unique selectivity. On the basis of the biochemical data, binding of NMM to Tel22 does not rely on relatively nonspecific electrostatic interactions, which characterize most canonical GQ ligands, but rather it is hydrophobic in nature. The structural features observed in the NMM–Tel22 complex described here will serve as guidelines for developing new quadruplex ligands that have excellent affinity and precisely defined selectivity.