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Hydrophobic mutations in buried polar residues enhance HIV-1 gp41 N-terminal heptad repeatC-terminal heptad repeat interactions and C-peptides anti-HIV activity

Zheng, Baohuaa,b; Wang, Kunb; Lu, Luc; Yu, Feid; Cheng, Maoshenga; Jiang, Shiboc,d,*; Liu, Kelianga,b,*; Cai, Lifengb,*

doi: 10.1097/QAD.0000000000000255
Basic Science

Objective: To investigate the effect of mutations in a highly conserved buried polar area on the function of HIV-1 gp41.

Design: During HIV-1 entry, a six helical bundle (6-HB) formation between the C-terminal and N-terminal heptad repeat (CHR and NHR) of gp41 provides energy for virus cell membrane fusion. In 6-HB, residues at a and d (a–d) positions of CHR directly interact with NHR and are buried. They are considered critical residues for 6-HB stability and for anti-HIV-1 activity of CHR-derived peptides (C-peptides). Most of a–d residues in CHR are hydrophobic, as buried hydrophobic residues facilitate protein stability. However, HIV-1 gp41 CHR contains a highly conserved polar area with four successive buried ad polar residues: S649/Q652/N656/E659. We mutated these buried polar residues to hydrophobic residues, either Leu or Ile, and studied its effect on the gp41 NHR–CHR interactions and anti-HIV activities of the C-peptides.

Methods: We measured the C-peptide mutants’ ability to form 6-HB with NHR, thermal stability of the 6-HBs and C-peptides’ inhibitory activity against both T20-sensitive and resistant HIV-1 strains.

Results: All the mutated C-peptides retained their ability to form stable 6-HB with NHR and strongly inhibited HIV-1 replication. Strikingly, S649L and E659I mutations endow C-peptide with a significantly enhanced activity against T20-resistant HIV-1 strains.

Conclusion: The highly conserved buried ad polar residues in HIV-1 gp41 CHR can be mutated as a means of developing new fusion inhibitors against drug-resistant HIV-1 strains. The concept can also be utilized to design fusion inhibitors against other viruses with similar mechanisms.

aMOE Key Laboratory of Structure-Based Drug Design & Discovery, Shenyang Pharmaceutical University, Shenyang

bDepartment of Medicinal Chemistry, Beijing Institute of Pharmacology & Toxicology, Beijing

cKey Laboratory of Medical Molecular Virology of MOE/MOH, Shanghai Medical College, Fudan University, Shanghai, China

dLindsley F. Kimball Research Institute, New York Blood Center, New York, New York, USA.

*Lifeng Cai, Keliang Liu and Shibo Jiang contributed equally to this work.

Correspondence to Lifeng Cai, PhD, Beijing Institute of Pharmacology & Toxicology, 27 Taiping Road, Haidian District, Beijing 100850, China. Tel: +86 10 66930640; fax: +86 10 68211656; e-mail:

Received 11 November, 2013

Revised 23 January, 2014

Accepted 5 February, 2014

© 2014 Lippincott Williams & Wilkins, Inc.