Biology Reference
In-Depth Information
DISCUSSION
Emerging and re-emerging infectious diseases remain to be one of the major causes
of death worldwide. The current outbreak of avian influenza viruses is a major global
concern due to the increasing number of fatalities among the poultry as well as human
cases. Its highly mutative nature makes the current antiviral drugs not very effective.
Therefore, there has been a constant need for broad-spectrum antiviral drugs against
the currently circulating human as well as avian strains.
In this study, a phage displayed peptide library was used to select anti-viral pep-
tides against the AIV H9N2. At the end of biopanning, four different peptide sequenc-
es were identifi ed. Matching of these peptide sequences with protein sequences in the
protein data banks (Swiss Prot and NCBI) showed no signifi cant homology with any
protein sequences. It is possible that these peptides might mimic a discontinuous bind-
ing site in which amino acids are brought from different positions of a protein to form
an essential contact area with the virion [23, 24]. The lack of antiviral activity by the
control peptide as well as the wild type phage suggest that the antiviral property of the
peptides is specifi c to those peptides and neither a general property of any oligomeric
peptide or wild type M13 bacteriophages nor based on charge or hydrophobic interac-
tions. The peptide phage competition assay proved that the peptide displayed on the
phage surface not the other parts of the phage binds to the virus.
Among the four different fusion phages isolated from the phage display library, the
phage displaying the sequence NDFRSKT was selected for further analysis as it repre-
sented highest number of clones in the fi nal round of biopanning. Besides, the peptides
LPYAAKH, ILGDKVG, and QHSTKWF showed negligible or no anti-viral activity
(data not shown); therefore, no further analyses on these peptides were carried out.
The
in ovo
model has been previously employed successfully by our group
Ramanujam et al. [25] and Song et al. [26] to study the inhibitory effect of anti-viral
molecules against the Newcastle disease virus and infl uenza virus respectively. There-
fore, the antiviral activity of the synthetic peptides and the fusion phages themselves
(or simply denoted as inhibitory peptides hereafter) were investigated in embryonated
chicken eggs. All the peptides showed good anti-viral properties against AIV and in-
terestingly there was no signifi cant anti-viral effect found against NDV strain AF2240.
Pre-treatment with the peptides or fusion phages reduced the AIV titre manifold (from
two fold to six fold based on the type of peptide and number of days of treatment) in
the infected allantoic fl uid. But the post-infection treatment failed to protect the em-
bryo (data not shown). However, it should be noted that the peptide was injected only
once in the study and besides, the amino acids of the peptide were of L-isomers which
are more prone to protease degradation inside the allantoic cavity.
Nevertheless, both cyclic and linear forms of peptides as well as the fusion phages
proved their worth as antiviral molecules in varied potential levels. Among them, the
cyclic peptide possessing the sequence CNDFRSKTC showed higher antiviral proper-
ties. The reason maybe its small size (only 9 amino acids in length for cyclic peptide)
which helps its easy access to the respective binding site on the target molecule. More-
over, the cyclic peptides possess a stable structure due to the disulfi de bond formed
between the fl anked cysteine residues which help to attain a stable interaction at a
