Journal
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
Volume 111, Issue 51, Pages 18207-18212Publisher
NATL ACAD SCIENCES
DOI: 10.1073/pnas.1413592112
Keywords
fusion inhibitor; HIV-1 neutralization; X-ray crystallography; drug design; coiled coil
Categories
Funding
- European Union Seventh Framework Programme [HEALTH-F3-2007-201038]
- Andalusia Regional Government [Incentivos-BOJA-5-1-2008, P09-CVI-5063]
- Spanish Ministry of Economy and Competitiveness [BIO2012-39922-C02-01/02, BIO2008-00750-E]
- European Fund for Regional Development (FEDER) (EU)
- European Synchrotron Radiation Facility block allocation group (BAG) [MX-1406, MX1541]
- Spanish Synchrotron Radiation Facility ALBA [2012010072, 2012100378]
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During HIV-1 fusion to the host cell membrane, the N-terminal heptad repeat (NHR) and the C-terminal heptad repeat (CHR) of the envelope subunit gp41 become transiently exposed and accessible to fusion inhibitors or Abs. In this process, the NHR region adopts a trimeric coiled-coil conformation that can be a target for therapeutic intervention. Here, we present an approach to rationally design single-chain protein constructs that mimic the NHR coiled-coil surface. The proteins were built by connecting with short loops two parallel NHR helices and an antiparallel one with the inverse sequence followed by engineering of stabilizing interactions. The constructs were expressed in Escherichia coli, purified with high yield, and folded as highly stable helical coiled coils. The crystal structure of one of the constructs confirmed the predicted fold and its ability to accurately mimic an exposed gp41 NHR surface. These single-chain proteins bound to synthetic CHR peptides with very high affinity, and furthermore, they showed broad inhibitory activity of HIV-1 fusion on various pseudoviruses and primary isolates.
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