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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

CO ligand migration along possible pathways in Mt-trHbN.(A) Schematic representations of the residues involved in the heme distal site and tunnels, the two tunnels and cavities estimated with ILS for the wild type form. (B) Free energy profiles over STG8 and (C) LT along the connection between solvent, (trHb : CO)2 and (trHb : CO)1 cavities for wild type (green), VG8F (orange) and VG8W (violet) mutant forms are shown. Circles represent calculated free energy values with the ILS method and lines correspond to a fitting estimation of these calculated values. The x coordinate represents the Fe-CO distance along the pathways.
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f4: CO ligand migration along possible pathways in Mt-trHbN.(A) Schematic representations of the residues involved in the heme distal site and tunnels, the two tunnels and cavities estimated with ILS for the wild type form. (B) Free energy profiles over STG8 and (C) LT along the connection between solvent, (trHb : CO)2 and (trHb : CO)1 cavities for wild type (green), VG8F (orange) and VG8W (violet) mutant forms are shown. Circles represent calculated free energy values with the ILS method and lines correspond to a fitting estimation of these calculated values. The x coordinate represents the Fe-CO distance along the pathways.

Mentions: The wild type Mt-trHbN presents two tunnels available for ligand migration, the LT and the STG8 (Figure 4A). On the one hand, the LT connects three internal cavities: (trHb : CO)1, (trHb : CO)2 and (trHb : CO)3. The STG8, on the other hand, has only the distal site cavity, (trHb : CO)1, which is directly connected to the solvent. The VG8F mutant conserves both tunnels, although they are constrained compared to those in the wild type. In the VG8W case, however, the energy profiles suggest a completely blocked STG8 and a LT for which the accessibility to the iron heme is partially reduced.


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)

CO ligand migration along possible pathways in Mt-trHbN.(A) Schematic representations of the residues involved in the heme distal site and tunnels, the two tunnels and cavities estimated with ILS for the wild type form. (B) Free energy profiles over STG8 and (C) LT along the connection between solvent, (trHb : CO)2 and (trHb : CO)1 cavities for wild type (green), VG8F (orange) and VG8W (violet) mutant forms are shown. Circles represent calculated free energy values with the ILS method and lines correspond to a fitting estimation of these calculated values. The x coordinate represents the Fe-CO distance along the pathways.
© Copyright Policy - open-access
Related In: Results  -  Collection

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

f4: CO ligand migration along possible pathways in Mt-trHbN.(A) Schematic representations of the residues involved in the heme distal site and tunnels, the two tunnels and cavities estimated with ILS for the wild type form. (B) Free energy profiles over STG8 and (C) LT along the connection between solvent, (trHb : CO)2 and (trHb : CO)1 cavities for wild type (green), VG8F (orange) and VG8W (violet) mutant forms are shown. Circles represent calculated free energy values with the ILS method and lines correspond to a fitting estimation of these calculated values. The x coordinate represents the Fe-CO distance along the pathways.
Mentions: The wild type Mt-trHbN presents two tunnels available for ligand migration, the LT and the STG8 (Figure 4A). On the one hand, the LT connects three internal cavities: (trHb : CO)1, (trHb : CO)2 and (trHb : CO)3. The STG8, on the other hand, has only the distal site cavity, (trHb : CO)1, which is directly connected to the solvent. The VG8F mutant conserves both tunnels, although they are constrained compared to those in the wild type. In the VG8W case, however, the energy profiles suggest a completely blocked STG8 and a LT for which the accessibility to the iron heme is partially reduced.

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