19
to be less susceptible toward drugs targeting the single system. However, the relation
between siderophore biosynthesis and virulence is complex, and thus, this architecture
networking needs to be explored more in terms of its understanding at cellular level.
2.3.1 Screening Through Biomonitor Organism
A generic screen was identified for finding biosynthetic inhibitors of siderophores
which proves to be beneficial in terms of cost and effectiveness. The opportunistic
fungal pathogen A. fumigatus is widely used as biomonitor organism (generic
screen) for screening purpose. This protocol is divided into two steps wherein at
first step, A. fumigatus is grown in either iron-deficient or iron-rich medium.
Compounds which show sluggish growth of pathogen in iron-limiting medium but
allow growth in iron-rich medium are taken forward for further evaluation. In the
second step, the fungi are grown again in iron-limiting medium, and those compounds which have shown limited production of siderophore which is confirmed
and quantified by colorimetric assay were considered as potential candidates. In
presence of ferric ions, the Fe(III)-siderophore complex displays red color which
shows reduction in intensity in presence of blockers of siderophore biosynthetic
pathway (Pinto and Moore 2009).
2.3.2 Screening of Compounds by Using Structural Similarity
Approach
This is yet another approach based on identification of siderophore inhibitors
which resemble siderophores more closely in terms of their structural features.
This strategy as of now is more utilized in targeting bacterial pathogenesis of
Mycobacterium tuberculosis and Yersinia pestis. Mycobactin and yersiniabactin
are the corresponding siderophores produced by these organisms (Stirrett et al.
2008). The pharmacophoric features of the compounds were identified which
cause inhibition of siderophore production, and thus, the pharmacophore can be
used for screening the library of compounds which can act as structural mimics of
siderophore. Based on EC 50 values and interference in bacterial growth, the
screened compounds were categorized as bacteriostatic and bactericidal.
Ravichandiran et al. conducted docking- based studies to screen plant-based inhibitors of SdiA. The top five hits were assessed for their binding affinity, and in vitro
assay was performed where compound 7-(1-bromoethyl)-3,3-dimethyl-bicyclo
[4.1.0] heptan-2 was found to interact with PHE 63, TYR 67, TRP 71, and VAL 86
amino acid residues in the active site of SidA where it competes with the natural
ligand (Ravichandiran et al. 2012).
2 Inhibition of Siderophores in Blocking Fungal Infection
to be less susceptible toward drugs targeting the single system. However, the relation
between siderophore biosynthesis and virulence is complex, and thus, this architecture
networking needs to be explored more in terms of its understanding at cellular level.
2.3.1 Screening Through Biomonitor Organism
A generic screen was identified for finding biosynthetic inhibitors of siderophores
which proves to be beneficial in terms of cost and effectiveness. The opportunistic
fungal pathogen A. fumigatus is widely used as biomonitor organism (generic
screen) for screening purpose. This protocol is divided into two steps wherein at
first step, A. fumigatus is grown in either iron-deficient or iron-rich medium.
Compounds which show sluggish growth of pathogen in iron-limiting medium but
allow growth in iron-rich medium are taken forward for further evaluation. In the
second step, the fungi are grown again in iron-limiting medium, and those compounds which have shown limited production of siderophore which is confirmed
and quantified by colorimetric assay were considered as potential candidates. In
presence of ferric ions, the Fe(III)-siderophore complex displays red color which
shows reduction in intensity in presence of blockers of siderophore biosynthetic
pathway (Pinto and Moore 2009).
2.3.2 Screening of Compounds by Using Structural Similarity
Approach
This is yet another approach based on identification of siderophore inhibitors
which resemble siderophores more closely in terms of their structural features.
This strategy as of now is more utilized in targeting bacterial pathogenesis of
Mycobacterium tuberculosis and Yersinia pestis. Mycobactin and yersiniabactin
are the corresponding siderophores produced by these organisms (Stirrett et al.
2008). The pharmacophoric features of the compounds were identified which
cause inhibition of siderophore production, and thus, the pharmacophore can be
used for screening the library of compounds which can act as structural mimics of
siderophore. Based on EC 50 values and interference in bacterial growth, the
screened compounds were categorized as bacteriostatic and bactericidal.
Ravichandiran et al. conducted docking- based studies to screen plant-based inhibitors of SdiA. The top five hits were assessed for their binding affinity, and in vitro
assay was performed where compound 7-(1-bromoethyl)-3,3-dimethyl-bicyclo
[4.1.0] heptan-2 was found to interact with PHE 63, TYR 67, TRP 71, and VAL 86
amino acid residues in the active site of SidA where it competes with the natural
ligand (Ravichandiran et al. 2012).
2 Inhibition of Siderophores in Blocking Fungal Infection