Abstract | By means of combined MD and QM/MM calculation we show that this conformation positions the NH backbone towards the sulfenic acid and promotes the reaction to yield the sulfenyl amide intermediate, in one step with the concomitant release of a water molecule . |
Protein structure selection, search parameters and Cys environment characterization | Each protein was then immersed in a truncated octahedral box of TIP3P water consisting in 8,776 water molecules , which corresponds to a 10 A distance between the protein surface and the box boundary [48]. |
QM/MM methods | The first step consists of the breakage of the S-OH bond (with OH leaving as a water molecule after proton transfer from H20 or H3O+) and the formation of the SN bond using the following reaction coordinate |
QM/MM methods | The second step involves the transfer of the amide proton to a water molecule , regenerating the H20 or H3O+molecule. |
QM/MM methods | In the case of the peptide, the QM system comprises the whole peptide (ACE-Cys-SOH-NME), H3O+ molecule and the closest 9 water molecules to the system. |
Results | To test this idea, we included in the QM system 10 water molecules and explicitly promoted proton transfer from the solvent to the S-OH group. |
Results | 4 shows a completely broken SO bond, a well formed water molecule and the S and N atoms quite close at a distance of 1.93 angstroms. |
Results | We observe only an increase in the OS atom that is due to its transfer from the sulfenic molecule to form a water molecule . |
Important interactions stabilizing the transition state | Apart from the electrostatic effects of the metal cation and hydrogen bonding with the water molecules coordinated to this ion, the leaving group is additionally forming a strong hydrogen bond with Tyr367, a hydrogen bond with Arg362 and another, relatively weak hydrogen bond with the amidic hydrogen of the acceptor threonine. |
Methods | Furthermore, water molecules present in the active site were manually rotated to create a network of hydrogen bonds where possible. |
Methods | Finally, 12 highly conserved residues interacting with the substrates were included (Table 2) as well as six water molecules that were well defined in the crystal structure (B-factors below 20). |
Methods | Three of these water molecules are located close to the metal ion with one of them directly serving as a ligand and the other two forming a hydrogen bond network between the first water molecule and neighboring active site residues. |
Inner Membrane Model | The DAB residues were observed to drag some water molecules into the hydrophobic bilayer core (Fig 3); on average less than one water molecule was present in the PMB1 free membrane, compared to an average of seven being present during the last microsecond of simulation. |
Inner Membrane Model | Moreover, when we studied the radial distribution of solvent around DAB residues, despite being fully inserted within the non-polar acyl tail region, an average of three water molecules were still present within 0.35 nm of the DAB nitrogen atoms (Fig 3). |
Inner Membrane Model | This is further supported by analysis of the mass density profile (Fig 3) that reveals increased water penetration, with water molecules reaching the hydrophobic core of the membrane as discussed above. |
Lipid A Outer Membrane Model | Interaction of the DAB residues with the membrane surface during aggregation caused water molecules to move towards the membrane. |
Re LPS Outer Membrane Model | Radial distribution functions (S4 Fig) showed an increased degree of solvation in the sugar group area of the membrane, such that on average 20 water molecules were present in this region after 2 pi-croseconds, compared to ~12 molecules prior to PMB1 binding. |