Anti-infective activity of apolipoprotein domain derived peptides in vitro: identification of novel antimicrobial peptides related to apolipoprotein B with anti-HIV activity

Abstract

Background: Previous reports have shown that peptides derived from the apolipoprotein E receptor binding region and the amphipathic alpha-helical domains of apolipoprotein AI have broad anti-infective activity and antiviral activity respectively. Lipoproteins and viruses share a similar cell biological niche, being of overlapping size and displaying similar interactions with mammalian cells and receptors, which may have led to other antiviral sequences arising within apolipoproteins, in addition to those previously reported. We therefore designed a series of peptides based around either apolipoprotein receptor binding regions, or amphipathic alpha-helical domains, and tested these for antiviral and antibacterial activity.

Results: Of the nineteen new peptides tested, seven showed some anti-infective activity, with two of these being derived from two apolipoproteins not previously used to derive anti-infective sequences. Apolipoprotein J (151-170) - based on a predicted amphipathic alpha-helical domain from apolipoprotein J - had measurable anti-HSV1 activity, as did apolipoprotein B (3359-3367) dp (apoBdp), the latter being derived from the LDL receptor binding domain B of apolipoprotein B. The more active peptide - apoBdp - showed similarity to the previously reported apoE derived anti-infective peptide, and further modification of the apoBdp sequence to align the charge distribution more closely to that of apoEdp or to introduce aromatic residues resulted in increased breadth and potency of activity. The most active peptide of this type showed similar potent anti-HIV activity, comparable to that we previously reported for the apoE derived peptide apoEdpL-W.

Conclusions: These data suggest that further antimicrobial peptides may be obtained using human apolipoprotein sequences, selecting regions with either amphipathic alpha-helical structure, or those linked to receptor-binding regions. The finding that an amphipathic alpha-helical region of apolipoprotein J has antiviral activity comparable with that for the previously reported apolipoprotein AI derived peptide 18A, suggests that full-length apolipoprotein J may also have such activity, as has been reported for full-length apolipoprotein AI. Although the strength of the anti-infective activity of the sequences identified was limited, this could be increased substantially by developing related mutant peptides. Indeed the apolipoprotein B-derived peptide mutants uncovered by the present study may have utility as HIV therapeutics or microbicides.

Figure 1

Anti-infective activity of alternative apoE(141-149) tandem repeat peptides derived from the human, murine and bovine apoE sequences. HSV1 infectivity after treatment with peptides derived from the human, mouse and bovine apoE141-149 sequence, shown relative to that for untreated controls. Human apoEdp had activity significantly different from untreated wells (P < 0.001) unlike the murine and bovine equivalent peptides. Typical data are shown here; bars indicating standard error.

Figure 2

Edmondson helical-wheel diagrams showing charge distribution of apoE and apoB derived peptides in α-helical conformation. Charged residues in apoEdp, apoBdp and apoBdpL/R do not show amphipathic distribution (like for example the lytic peptide RLLR5), though are arranged in a bipolar asymmetric pattern which is more pronounced in apoEdp and apoBdpL/R.

Figure 3

Antiviral and antibacterial activity of apoBdprelated peptides. (A) A peptide consisting of a tandem repeat peptide of the naturally occurring apoB3359-3367 sequence has limited antiviral activity which is enhanced in the apoBdpL/R modified sequence, though (B) antibacterial activity was not measurable for both. (C) ApoBdpL/R has comparable antiviral activity to apoEdp though (D) does not share its strong antibacterial activity. (E) The Trp-substituted apoBdpL/R-W peptide has stronger antiviral activity and (F) shows similar antibacterial activity. Values shown are HSV1 infectivity after plaque reduction assay in Vero cells or turbidity of Pseudomonas aeruginosa; typical data are shown; bars indicate standard error.

Figure 4

Reduction of antiviral and antibacterial activity of apoEdpL/R-W peptides after alteration of Arg-Lys-Arg repeated motif. Substitution of the repeated Arg-Lys-Arg motif within apoBdpL/R-W for Arg-Arg-Arg (apoBdpL/R-WR) or Lys-Lys-Lys (apoBdpL/R-WK) reduces (A) antiviral activity or (B) antibacterial activity. Values shown are HSV1 infectivity after plaque reduction assay in Vero cells or turbidity of Pseudomonas aeruginosa; typical data are shown; bars indicate standard error.

Figure 5

Anti-HIV activity of the apolipoprotein B derived peptide apoBdpL/R-W. (A) ApoBdpL/R-W has potent activity against a number of HIV strains, including both CXCR4 and CCR5 co-receptor using strains, similar to that we previously reported for apoEdpL-W. Experiments were performed by treating TZMbl cells with peptide prior to introducing the CXCR4 using HIV strain HIV NL4.3, the CCR5 using HIV strain BaL, and the dual co-receptor using HIV strain 89.6. Values show level of HIV infection (in RLU) relative to untreated control; bars indicate standard errors. No effect was seen on growth of TZMbl cells over the concentration range tested against HIV. ApoBdpL-W does not inhibit VSV-g infection (B) or cause haemolysis (C) suggesting its activity is relatively selective.