Molecular model of hemoglobin N from Mycobacterium tuberculosis bound to lipid bilayers: a combined spectroscopic and computational study.
A singular aspect of the 2-on-2 hemoglobin structures of groups I and II is the presence of tunnels linking the protein surface to the distal heme pocket, supporting the storage and the diffusion of small apolar ligands to/from the buried active site. As the solubility of apolar ligands is greater in biological membranes than in solution, the association of these proteins with biological membranes may improve the efficiency of ligand capture. As very little is known on this subject, we have investigated the interactions between hemoglobin N (HbN), a group I 2-on-2 hemoglobin from the pathogenic Mycobacterium tuberculosis (Mtb), and biological membranes using both experimental techniques and MD simulations. HbN has a potent nitric oxide dioxygenase activity (HbN-Fe²⁺-O₂ + •NO + H₂O → HbN-Fe³⁺-OH₂ + NO₃⁻) that is thought to protect the aerobic respiration of Mtb from inhibition by •NO. Three different membrane compositions were chosen for the studies, representative of the mycobacterial plasma membrane and the mammalian cell membranes. Both the experimental and the modeling results agreed with each other and allow for a detailed molecular description of HbN in association with membranes of different compositions. The results indicated that HbN is a peripheral protein, and the association with the membranes occurred via the pre-A, G, and H helices. In addition, HbN would be allowed to modulate the binding to the membranes via electrostatic interactions between the lipid membranes and the Asp100 residue. In its membrane-bound form the short tunnel of HbN is oriented toward the membrane interior and the other tunnels point toward the solvent. Such protein orientation would facilitate the uptake of nonpolar substrates from the membrane and the release of products to the solvent. It is interesting to note that the pre-A, G, and H helices are conserved among HbN from a few other Mycobacteria.
‡Department of Biochemistry, Microbiology and Bioinformatics, Université Laval, Québec, Québec, Canada.
Jean-François
JF
Rhéault
Ève
È
Gagné
‡Department of Biochemistry, Microbiology and Bioinformatics, Université Laval, Québec, Québec, Canada.
Michel
M
Guertin
⊥Centre for Research in Molecular Modeling (CERMM), Concordia University, Montréal, Québec, Canada.
Guillaume
G
Lamoureux
Michèle
M
Auger
‡Department of Biochemistry, Microbiology and Bioinformatics, Université Laval, Québec, Québec, Canada.
Patrick
P
Lagüe
D000595:Amino Acid Sequence; D001224:Aspartic Acid / Q000737:chemistry; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism; D002308:Cardiolipins / Q000737:chemistry / Q000378:metabolism; D002942:Circular Dichroism; D017124:Conserved Sequence; D030562:Databases, Protein; D008051:Lipid Bilayers / Q000737:chemistry* / Q000378:metabolism; D008958:Models, Molecular; D056004:Molecular Dynamics Simulation; D009169:Mycobacterium tuberculosis / Q000201:enzymology*; D019906:Nuclear Magnetic Resonance, Biomolecular; D010105:Oxygenases / Q000737:chemistry* / Q000378:metabolism; D010713:Phosphatidylcholines / Q000737:chemistry / Q000378:metabolism; D010714:Phosphatidylethanolamines / Q000737:chemistry / Q000378:metabolism; D011487:Protein Conformation; D017550:Spectroscopy, Fourier Transform Infrared; D055672:Static Electricity; D054793:Truncated Hemoglobins / Q000737:chemistry* / Q000378:metabolism
C519576:1,1',2,2'-tetraoleoylcardiolipin; D001426:Bacterial Proteins; D002308:Cardiolipins; C102539:GlbN protein, bacteria; D008051:Lipid Bilayers; D010713:Phosphatidylcholines; D010714:Phosphatidylethanolamines; D054793:Truncated Hemoglobins; C094877:dioleoyl phosphatidylethanolamine; D001224:Aspartic Acid; D010105:Oxygenases; C114415:nitric oxide dioxygenase; C017251:1,2-oleoylphosphatidylcholine