Abstract
The interface between the HIV-1 gp120 envelope glycoprotein and the CD4 receptor contains an unusual interfacial cavity, the "Phe43 cavity", which CD4-mimetic miniproteins with nonnatural extensions can potentially utilize to enhance their neutralization of HIV-1. Here, we report cocrystal structures of HIV-1 gp120 with miniproteins M48U1 and M48U7, which insert cyclohexylmethoxy and 5-hydroxypentylmethoxy extensions, respectively, into the Phe43 cavity. Both inserts displayed flexibility and hydrophobic interactions, but the M48U1 insert showed better shape complementarity with the Phe43 cavity than the M48U7 insert. Subtle alteration in the gp120 conformation played a substantial role in optimizing fit. With M48U1, these translated into a YU2-gp120 affinity of 0.015 nM and neutralization of all 180 circulating HIV-1 strains tested, except clade-A/E isolates with noncanonical Phe43 cavities. Ligand chemistry, shape complementarity, surface burial, and gp120 conformation act in concert to modulate binding of ligands to the gp120-Phe43 cavity and, when optimized, can effect near-pan-neutralization of HIV-1.