Wild type NPC2 with bovine Lys-29 and Arg-32 replaced by mouse Asn-29 and Lys-32, respectively. The side chains of all mutants made in this study are shown as sticks with functional coloring (green, carbon; white, hydrogen; blue, nitrogen; red, oxygen) except for Val-64. Val-64 along with Tyr-100, Pro-101, Ile-103, Val-59, and Phe-66 form the outer rim of the cholesterol binding pocket as described in Fig. 6 of Xu et al. (8). The rim, with the full residues shown as sticks, is shown in magenta. This orientation shows the “front” of the protein.