L-RAR systems. Comparison of active site residues of WT and E271Q systems using the representative structures obtained from the simulations revealed the structural changes led to the differences in the catalytic activity of the enzyme ( Figure 10 ). The overlaid WT and E271Q structures have shown the difference in the locations of catalytically important Zn 2+ ion in the catalytic center ( Figure 10A ). The uncharged nature of the mutant residue Q271 the metal ion has lost the important interaction and moved far away from its original location. The distance between the Zn 2+ ion and the oxygen atom of the N-terminal peptide bond was so high compared to the WT system. Thus the catalytic aminopeptidase reaction becomes impossible in E271Q system. The helix containing Y383 was extended during the simulation E271Q system moving Y383 backward from the active site. This movement of Y383 has formed p-cation and a hydrogen bond interactions with the C-terminal part of RAR whereas in WT system this residue has formed a hydrogen bond interaction with the N-terminal amino group. The other tyrosine residue Y378, which was found to be guiding the substrate along with the carboxylate recognition site residues, has formed hydrogen bond with the carboxylate of RAR. The binding mode of RAR observed in E271Q system has shown only weak hydrogen bonding interactions with R563 as it was moving away from it. The helix formed by K565-A575 residues was shortened slightly leaving K565, one of the carboxylate recognition residues, as a part of loop making it more flexible. But this residue has maintained hydrogen bond interactions with the peptide substrate through its carboxylate group. This change observed in helix containing K565 is different from that of L-LTA4 systems. The K565 is the key residue that assists the other positively charged residue R563 to maintain the proper alignment of RAR in the active site whereas in LTA4 binding K565 is not required to assist R563 residue [27] . Moreover, the absence of negatively charged E271 residue also played a major role in observed loss of catalytic activity. The R563A mutation has caused a high fluctuation of E271 that moved far from the N-terminal amino group of RAR and makes it impossible to act as N-terminal recognition site ( Figure 10B ). The interacting distance between Zn 2+ and E271 was maintained in this mutation. The helix extension was observed near Y383 as displayed in E271Q system and this changed the flexibility of Y383 in the active site. The other tyrosine residue Y378 was highly fluctuating in this mutant system compared to any other systems and formed strong p-cation interaction than it is in WT system. The C-terminal part of RAR substrate has slightly went back as it missed the strong interactions from R563 but the alignment was almost maintained as observed in WT system except the side Cterminal side chain of RAR. Interestingly, K565 has taken the location of R563 in this mutant system to maintain the hydrogen bonds with the carboxylate of RAR and there was no change observed in the K565-A575 helix as observed in E271Q system. The K565A system also has shown some structural changes that were not observed in other L-RAR systems ( Figure 10C ). The short helix (V306-N308) that has shown structural changes in L-LTA4 system disappeared in K565A system and the F314 helix was extended including W311-F314 residues. These changes were observed in other L-RAR systems. The extension of helix has drawn F314 residue back from the active site center. A folded binding mode of RAR was observed in K565 system much different from that of WT and other mutant L-RAR systems. Though R563 is present, the carboxylate group of RAR has moved back from its original position and formed p-interactions with Y378 and completely lost interactions with R563. This observation completely correlates with the observed activity and the reported statement that K565 assists R563 to act as carboxylate recognition site in aligning the substrate along the catalytic elements of the enzyme. The overlay of all L-RAR systems revealed that along with E271, R563, K565 residues, Y378 and Y383 were also important in keeping the peptide substrate aligned within the active site ( Figure 10D ). As observed in L-LTA4 systems, Y378 residue has acted as a baffle to control the binding modes of the substrates. This part of the study has documented various structural changes explaining the differences in activities between WT and mutated forms of the enzyme bound to its two different substrates which were not determined by the X-ray crystallography so far (Table S1 ).