Fig 1.
A schematic representation of nitrate transport.
(A) The TMD proteins of the ABC-nitrate transporter, periplasmic subunit nrtA scavenges nitrate of the solution and transports across the lipid bilayer of the membrane through nrtB pore. When nitrate level rises in the cell, ABC protein, NrtC binds to nitrate and acts as negative regulator by inhibiting nitrate uptake, while nrtD binds and hydrolyzes ATP to induce conformational changes in TMDs (nrtAB). Extra cytoplasmic helices running parallel to the cytoplasmic membrane provide contact to TMDs proteins. (B) Interaction of GB with nrtACD proteins.
Fig 2.
Nitrate uptake in Anabaena 7120, WT and ApGSMT-DMT transformant.
Exponential phase cyanobacterial cells growing on N2 in the presence and absence of 0.1 M NaCl were collected and suspended in their specific media containing varied concentrations (40–500 μ) of nitrate. Uptake was performed under standard growth conditions in light (70 μE m2 s1). Samples were withdrawn at 10 min of incubation and measured nitrate depletion in the incubation mixture. Values represent means and SDs of triplicate experiments. (A) Initial rate of nitrate transport at different substrate concentrations; (B) Hofstee plot (v versus v/[S]) to determine the kinetics.
Fig 3.
Phylogenetic tree of deduced nrtA protein.
The tree was constructed by PHYLIP programs using Seqboot, Protdist, Neighbour and Consense methods. Bootstraping (1000 times) was performed to obtain support values for each branch. Values are shown at the nodes of each branch point.
Fig 4.
Phylogenetic tree of deduced nrtC protein.
The tree was constructed by PHYLIP programs using Seqboot, Protdist, Neighbour and Consense methods. Bootstraping (1000 times) was performed to obtain support values for each branch. Values are shown at the nodes of each branch point.
Fig 5.
Phylogenetic tree of deduced nrtD protein.
The tree was constructed by PHYLIP programs using Seqboot, Protdist, Neighbour and Consense methods. Bootstraping (1000 times) was performed to obtain support values for each branch. Values are shown at the nodes of each branch point.
Fig 6.
Homology model of nrtACD proteins.
Models are presented as ribbon structure. Alpha helices are in red, beta sheets in cyan, turn in grey and coil in green color. (A) nrtA, (B) nrtC and (C) nrtD.
Fig 7.
RAMPAGE analysis of nrtACD proteins.
Values indicate number of the residues in favored, allowed, and outlier region. (A) nrtA, (B) nrtC and (C) nrtD.
Fig 8.
Ramachandran plot analysis of traget proteins nrtACD.
Red regions in the graph indicate the most allowed regions and yellow the allowed region. (A) nrtA, (B) nrtC and (C) nrtD.
Fig 9.
ERRAT measurement of nrtA protein.
Values indicate over all structure quality of the traget protein.
Fig 10.
ERRAT measurement of nrtC protein.
Values indicate over all structure quality of the traget protein.
Fig 11.
ERRAT measurement of nrtD protein.
Values indicate over all structure quality of the traget protein.
Fig 12.
ProSA analysis for nrtA (A), nrtC (B), nrtD (C) proteins.
Z-scores of the protein chains in PDB were determined by X-ray crystallography (light blue) and NMR spectroscopy (dark blue). The proSA web results indicated that protein structures had characteristic features of their native structures. The Z-score for all the three target proteins was highlighted as a large dot.
Table 1.
Similarity between template and target proteins nrtACD of ABC-transporter based on identities and positives of model proteins.
Fig 13.
Docking of ligands GB and nitrate with nrtA protein.
(A) Green color in the sphere indicates prominent active site with which the ligand interacted. (B) 3D level interaction.
Fig 14.
Docking of ligands GB and nitrate with target protein nrtC.
(A) Poses of docked complexes; green color in the sphere indicates prominent active site on which the ligand interacted. (B) 3D level interaction.
Fig 15.
Docking of ligands GB and nitrate with target protein nrtD.
(A) Poses of docked complexes; green color in sphere indicates prominent active site, on which the ligand interacted. (B) 3D level interaction.
Table 2.
The potential ligand binding catalytic active sites of target proteins (nrtACD) calculated with METAPOCKET server to perform molecular docking.
Table 3.
PatchDock server molecular docking results of GB and nitrate with ABC transporter proteins nrtACD.