Characterization of the steric defense of the HIV-1 gp41 N-trimer region.
During viral entry, HIV gp41 adopts a transient conformation called the "prehairpin intermediate" in which a highly conserved therapeutic target, the N-trimer, is exposed. Despite extensive discovery efforts, potent and broadly neutralizing antibodies that target the N-trimer are elusive. We previously demonstrated the N-trimer is protected by a steric block that prevents large proteins, such as antibodies, from accessing it. Here we further characterize the steric block and identify its source. To study the N-trimer steric accessibility, we produced two sets of C-peptide inhibitors (a potent inhibitor targeting the N-trimer) fused to cargo proteins of increasing size facing either the virus or cell side of the prehairpin intermediate. Both bulky inhibitor sets show a steric block, but the effect is more pronounced with virus-side cargo. Additionally, both sets maintain their potencies in a modified entry assay that removes possible sources of target cell steric hindrance. These results implicate a viral source, likely gp120, as the primary component of the steric block. In addition, we studied the steric accessibility of the "pocket" region of the N-trimer, a highly attractive drug and vaccine target. We demonstrated a pocket-specific antibody, D5, is more potent as an scFv than as a full-length IgG, suggesting the N-trimer steric restriction extends to the pocket. This characterization will facilitate the design of sterically restricted antigens that mimic the steric environment of the N-trimer in the prehairpin intermediate and are capable of inducing potent and broadly neutralizing antibodies that circumvent the N-trimer steric block.
Eckert
Debra M
DM
Department of Biochemistry, University of Utah School of Medicine, Salt Lake City, Utah 84112-5650, USA.
Shi
Yu
Y
Kim
Sunghwan
S
Welch
Brett D
BD
Kang
Eunchai
E
Poff
Emily S
ES
Kay
Michael S
MS
D015700:HIV Envelope Protein gp41 / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008561:Membrane Fusion; D010446:Peptide Fragments / Q000037:antagonists & inhibitors; D011485:Protein Binding; D011487:Protein Conformation; D054730:Protein Interaction Domains and Motifs; D055550:Protein Stability; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000494:pharmacology; D020349:Surface Plasmon Resonance
D015700:HIV Envelope Protein gp41; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins
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