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Ligand uptake in Mycobacterium tuberculosis truncated hemoglobins is controlled by both internal tunnels and active site water molecules.

Boron I, Bustamante JP, Davidge KS, Singh S, Bowman LA, Tinajero-Trejo M, Carballal S, Radi R, Poole RK, Dikshit K, Estrin DA, Marti MA, Boechi L - F1000Res (2015)

Bottom Line: In order to investigate the differences between these proteins, we performed experimental kinetic measurements, (•)NO decomposition, as well as molecular dynamics simulations of the wild type Mt-trHbN and two mutants, VG8F and VG8W.These mutations introduce modifications in both tunnel topologies and affect the incoming ligand capacity to displace retained water molecules at the active site.We found that a single mutation allows Mt-trHbN to acquire ligand migration rates comparable to those observed for Mt-trHbO, confirming that ligand migration is regulated by the internal tunnel architecture as well as by water molecules stabilized in the active site.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Química Biológica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Buenos Aires, C1428EGA, Argentina.

ABSTRACT
Mycobacterium tuberculosis, the causative agent of human tuberculosis, has two proteins belonging to the truncated hemoglobin (trHb) family. Mt-trHbN presents well-defined internal hydrophobic tunnels that allow O 2 and (•)NO to migrate easily from the solvent to the active site, whereas Mt-trHbO possesses tunnels that are partially blocked by a few bulky residues, particularly a tryptophan at position G8. Differential ligand migration rates allow Mt-trHbN to detoxify (•)NO, a crucial step for pathogen survival once under attack by the immune system, much more efficiently than Mt-trHbO. In order to investigate the differences between these proteins, we performed experimental kinetic measurements, (•)NO decomposition, as well as molecular dynamics simulations of the wild type Mt-trHbN and two mutants, VG8F and VG8W. These mutations introduce modifications in both tunnel topologies and affect the incoming ligand capacity to displace retained water molecules at the active site. We found that a single mutation allows Mt-trHbN to acquire ligand migration rates comparable to those observed for Mt-trHbO, confirming that ligand migration is regulated by the internal tunnel architecture as well as by water molecules stabilized in the active site.

No MeSH data available.


Related in: MedlinePlus

Schematic representations of the distal site of Mt-trHbN.(A) wild type, (B) VG8F and (C) VG8W forms showing, on the basis of MD simulations, the hydrogen-bond network (dotted lines) stabilizing a water molecule above the iron heme. The percentages depicted as insets in the figure correspond to active site water occupancy during MD simulation.
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f5: Schematic representations of the distal site of Mt-trHbN.(A) wild type, (B) VG8F and (C) VG8W forms showing, on the basis of MD simulations, the hydrogen-bond network (dotted lines) stabilizing a water molecule above the iron heme. The percentages depicted as insets in the figure correspond to active site water occupancy during MD simulation.

Mentions: The active site water molecules occupancy was computed for all three systems by performing 200 ns of MD simulations with explicit water molecules. In each case a water molecule was able to access the active site and was stabilized by the iron and the distal site residues (Figure 5). Specifically, in both wild type and VG8F Mt-trHbN a water molecule was present for approximately 40% of the length of the simulation (Figure 5A, 5B). The VG8W mutant active site, on the other hand, is occupied by water molecules in 80% of the simulation time, probably due to the hydrogen bonding capacity of W (Figure 5C).


Ligand uptake in Mycobacterium tuberculosis truncated hemoglobins is controlled by both internal tunnels and active site water molecules.

Boron I, Bustamante JP, Davidge KS, Singh S, Bowman LA, Tinajero-Trejo M, Carballal S, Radi R, Poole RK, Dikshit K, Estrin DA, Marti MA, Boechi L - F1000Res (2015)

Schematic representations of the distal site of Mt-trHbN.(A) wild type, (B) VG8F and (C) VG8W forms showing, on the basis of MD simulations, the hydrogen-bond network (dotted lines) stabilizing a water molecule above the iron heme. The percentages depicted as insets in the figure correspond to active site water occupancy during MD simulation.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4591903&req=5

f5: Schematic representations of the distal site of Mt-trHbN.(A) wild type, (B) VG8F and (C) VG8W forms showing, on the basis of MD simulations, the hydrogen-bond network (dotted lines) stabilizing a water molecule above the iron heme. The percentages depicted as insets in the figure correspond to active site water occupancy during MD simulation.
Mentions: The active site water molecules occupancy was computed for all three systems by performing 200 ns of MD simulations with explicit water molecules. In each case a water molecule was able to access the active site and was stabilized by the iron and the distal site residues (Figure 5). Specifically, in both wild type and VG8F Mt-trHbN a water molecule was present for approximately 40% of the length of the simulation (Figure 5A, 5B). The VG8W mutant active site, on the other hand, is occupied by water molecules in 80% of the simulation time, probably due to the hydrogen bonding capacity of W (Figure 5C).

Bottom Line: In order to investigate the differences between these proteins, we performed experimental kinetic measurements, (•)NO decomposition, as well as molecular dynamics simulations of the wild type Mt-trHbN and two mutants, VG8F and VG8W.These mutations introduce modifications in both tunnel topologies and affect the incoming ligand capacity to displace retained water molecules at the active site.We found that a single mutation allows Mt-trHbN to acquire ligand migration rates comparable to those observed for Mt-trHbO, confirming that ligand migration is regulated by the internal tunnel architecture as well as by water molecules stabilized in the active site.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Química Biológica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Buenos Aires, C1428EGA, Argentina.

ABSTRACT
Mycobacterium tuberculosis, the causative agent of human tuberculosis, has two proteins belonging to the truncated hemoglobin (trHb) family. Mt-trHbN presents well-defined internal hydrophobic tunnels that allow O 2 and (•)NO to migrate easily from the solvent to the active site, whereas Mt-trHbO possesses tunnels that are partially blocked by a few bulky residues, particularly a tryptophan at position G8. Differential ligand migration rates allow Mt-trHbN to detoxify (•)NO, a crucial step for pathogen survival once under attack by the immune system, much more efficiently than Mt-trHbO. In order to investigate the differences between these proteins, we performed experimental kinetic measurements, (•)NO decomposition, as well as molecular dynamics simulations of the wild type Mt-trHbN and two mutants, VG8F and VG8W. These mutations introduce modifications in both tunnel topologies and affect the incoming ligand capacity to displace retained water molecules at the active site. We found that a single mutation allows Mt-trHbN to acquire ligand migration rates comparable to those observed for Mt-trHbO, confirming that ligand migration is regulated by the internal tunnel architecture as well as by water molecules stabilized in the active site.

No MeSH data available.


Related in: MedlinePlus