15 research outputs found
The N‑Terminal T–T Motif of a Third-Generation HIV‑1 Fusion Inhibitor Is Not Required for Binding Affinity and Antiviral Activity
The
highlighted next-generation HIV-1 fusion inhibitor peptide <b>1</b> is capped by two threonines. Here, we generated peptide <b>2</b> by deleting the T–T motif and compared their structural and
antiviral properties. Significantly, two peptides showed similar helical
and oligomeric states in solution, comparable binding affinities to
the target, and no significant difference to inhibit HIV-1 fusion
and infection. Also, the T–T motif was not associated with
peptide <b>1</b> resistant mutations and its deletion did not
affect peptide <b>1</b> against enfuvirtide-resistant HIV-1
mutants. The redundancy of the T–T motif was further verified
by the model peptide C34 and short peptide inhibitors that mainly
target the gp41 pocket, suggesting that the N-terminal T–T
motif of peptide <b>1</b> could be removed or modified toward
the development of new anti-HIV-1 drugs. Consistently, our data have
verified that the M–T hook structure rather than the T–T
motif is an efficient strategy for short peptide fusion inhibitors
Inhibition of ABT on HIV-1 entry and replication.
<p>A. Inhibition of HIV-1<sub>NL4-3</sub> Env-pseudotyped virus in single-cycle assay that demonstrates the virus-cell membrane fusion. B. Inhibition of wild-type HIV-1<sub>NL4-3</sub> replication. ABT shows significantly higher potency than T20 in inhibiting HIV-1<sub>NL4-3</sub> entry and replication. The data were derived from the results of three independent experiments and are expressed as means ± standard deviations.</p
Effect of human serum on the anti-HIV activity of ABT.
<p>The human sera were freshly isolated from a HIV-seronegative healthy volunteer. The peptide was mixed with various concentrations (5, 10, 20 and 50%) of human sera freshly isolated from a HIV-seronegative healthy volunteer and incubated for 2 h at 37°C. The mixture was then diluted with a DMEM-based complete medium supplemented with 10% FCS and subjected to the single-cycle infection assay. The percent inhibition by ABT was calculated as described in the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0032599#s4" target="_blank">materials and method</a>. The data were derived from the results of three independent experiments and are expressed as means ± standard deviations.</p
Schematic illustration of HIV-1 gp41 and peptide fusion inhibitors.
<p>A. View of the gp41 functional regions. The residue numbers of each region correspond to their positions in gp160 of HIV-1<sub>HXB2</sub>. FP, fusion peptide; NHR, N-terminal heptad repeat; CHR, C-terminal heptad repeat; TM, transmembrane domain. B. Sequence of CHR-derived anti-HIV-1 peptides. ABT is engineered with three amino acids different from C34 (marked in bold). The 13th residue serine (S) of C34 was changed to lysine (K) which allows a single modification by 3-maleimidopropionic acid (MPA).</p
Inhibition of ABT on subtype A, B and C HIV-1 strains<sup>a</sup>.
a<p>The inhibitory activity of each peptide was determined in triplicate by a single-cycle infectivity assay. The data were derived from the results of three independent experiments and are expressed as means ± standard deviations.</p>b<p>Co-R: coreceptor use.</p
Biophysical characterization of ABT by CD spectroscopy.
<p>A. CD spectra of NHR and CHR-derived peptides and their complexes. B. Thermostability of the complex formed by N36 and ABT or C34. The unfolding temperature of each complex was scanned at 222 nm by CD spectroscopy, and their <i>Tm</i> values were calculated. The final concentration of each peptide in PBS is 1 µM.</p
Inhibition of ABT on CRF07_BC, CRF01_AE and B'HIV-1 variants<sup>a</sup>.
a<p>The inhibitory activity of each peptide was determined in triplicate by a single-cycle infectivity assay. The data were derived from the results of at least three independent experiments and are expressed as means ± standard deviations.</p>b<p>HIV-1 subtypes: B/C, CRF07_BC; A/E, CRF01_AE; B′, Tai B.</p>c<p>Co-R: coreceptor use.</p
Inhibition of ABT on 6-HB formation and cell membrane fusion.
<p>A. ABT and C34 can efficiently inhibit 6-HB formation in a dose-dependent manner, but T20 has no such effect. B. Inhibition of HIV-1<sub>HXB2</sub> Env-mediated cell-cell membrane fusion by ABT, C34 and T20. The data were derived from the results of three independent experiments and are expressed as means ± standard deviations.</p
Inhibition of ABT on T20-resistant HIV-1 variants<sup>a</sup>.
a<p>The data were derived from the results of three independent experiments and are expressed as means ± standard deviations.</p>b<p>HIV-1NL4-3-based pseudoviruses were constructed and used in the single-cycle infection assays except that L33S and I37V/V38T are infectious molecular clones of HIV-1NL4-3.</p>c<p>Both wild-type (WT) and D36G were used as reference viruses to calculate the resistance fold-changes.Shown in parentheses are based on the D36G as a reference.</p
Molecular docking models of m36.
<p><b>A</b>. Zdock score from gp120-m36 interaction simulations. <b>B.</b> E_Rock for the best poses from the result of ZDock. <b>C.</b> The interface sequence of the three top-score poses (575, 1734, 1955). The residues predicted by both Mapitope and molecular docking methods are underlined.</p