Theoretical Investigation of TrHbN Association to Biological Membranes

Abstract

TrHbN is a group I truncated hemoglobin from Mycobacterium tuberculosis (Mtb). TrHbN protects Mtb aerobic respiration from •NO inhibition and is thought to play pivotal roles in the persistence of mycobacterial infection. Its 3D structure is characterized by the presence of hydrophobic tunnels. Previous MD simulations revealed that three of theses tunnels are used by •NO to reach the active site. Despite the fact that trHbN is soluble and highly active in aqueous solvent, there is compelling evidence suggesting that trHbN is associated with biological membranes. We have investigated the association of trHbN with membranes differing in lipid compositions (DOPC, CL and CL:DOPE) using all-atom MD simulations.We found that trHbN penetrates the peripheral regions of the lipid bilayer for all the lipid compositions. Two portions of trHbN make a dominant contribution to the membrane binding: the N-terminal pre-A helix, and the G and H helices from the core of the protein. The pre-A helix has four charged residues that interact with the lipid phosphates, and several hydrophobic residues that insert under the lipid headgroups. The G and H helices have several hydrophobic residues in contact with the aliphatic chains. These residues were identified to form a large hydrophobic funnel-shape entrance of the Short tunnel (ST), previously shown to be preferentially used by •NO to reach the distal heme pocket. As a consequence, the ST has its entrance pointing towards the lipid bilayer, while the Long tunnel (LT) entrance is pointing towards the aqueous solvent. The orientation of trHbN differs slightly in presence of cardiolipins due to additional lipid phosphate-protein core interactions. These results suggest that the substrates reach the active site from the membrane interior using the ST and the products escape to the solvent using the LT.