• 2018-07
  • 2018-10
  • 2018-11
  • 2019-04
  • 2019-05
  • 2019-06
  • 2019-07
  • 2019-08
  • 2019-09
  • 2019-10
  • 2019-11
  • 2019-12
  • 2020-01
  • 2020-02
  • Six substrate recognition sites SRS SRS


    Six substrate recognition sites (SRS1-SRS6) have been identified at positions 102-123, 204-209, 238-244, 301-319, 374-382 and 483-492 (Table 2, underlined residues) based on Gotoh\'s proposal [30]. CYP3A163 displays the similar characteristic long sequences between helices F and G presenting two additional helices F′ and G’. The residues 202-207 and residues 218-225 represent helices F and F′ while residues 227-235 and residues 243–257 represent helices G′ and G. The residues between these helices do not exhibit a regular secondary structure, connect these helices and make the roof of the active site above haem group and are part of SRS2 and SRS3 respectively. This region has been marked as a membrane interaction domain with hydrophobic outer surfaces in human CYP3A4 [31] and is vital for ligand induced conformation VU 0155069 mg [12]. SRS4 is located on helix-I. Phenylalanine cluster in CYP3A4, scattered between the SRS1-SRS4 region and a role in conformation to close the roof of the active site above the haem group, comply the reported locations in CYP3A163 e.g. F-108, F-213, F-215, and F-241 except F-304 that is replaced by hydrophobic alanine (A-304). Mutations to the residues that are part of the roof above the active site e.g. L-210, F-213, and F-215, have confirmed the change in flexibility of the enzyme [31]. The residues between helices I and H have not been defined in the structures of CYP3A4 and are considered as a flexible region [32]. These residues mark the region of CYP3A163 indicating deletion of 1 residue (N-282) and 7 residues (NNGHVGN, 282-288) in birds and mammals respectively compared to CYP3A sequences in reptiles. These residues could be a characteristic feature of the reptilian CYP3A subfamily being common in the reptilian CYP3A sequences and possibly linked to substrate specificity. Helix-I known as the backbone of CYPs is extended across the entire molecule. The residues on helix-I near the haem group are important for the catalytic function [12], [33]. A conserved pentapeptide with a role in proton transfer XGXXT (AGYET, 312-316) is orientated parallel to the distal side of the haem group. The G-313 and T-316 impart a characteristic kink to the helix-I and are part of the molecular oxygen binding site. This motif strongly complies with its conservational characteristics, first residue being the alanine (A) whereas the one preceding T-316 is almost invariably acidic in character, being glutamic acid (E-315) [34]. The conserved P-332 marks the junction between helices I and J. Another absolute conserved tetrapeptide EXXR (ETLR, 374-377) in the K helix helps to stabilise the core structure [35], present on proximal side of the haem group towards the putative redox partners binding site [36]. The aromatic sequences TILKG (394-398) span the connection between β sheets and PEPEEFRPERF (416-426) is part of meander region (haem binding pocket) with P-423 being the highly conserved residue [37]. The tetrapeptide PERF (423-426) from this motif is the differentiating feature of eukaryotic CYPs involved in catalysis and substrate specificities [34]. The CYP3A4 structure manifest larger active site and a closely existing large solvent channel to facilitate oxidation of larger or multiple substrate. This solvent channel is formed by interaction among tyrosine (Y-53), aspartic acid (D-61), aspartic acid (D-76), arginine (R-106), arginine (R-372) and glutamic acid (E-374) [31]. These characteristic residues except the glutamic acid (E-374) that is replaced by less hydrophilic tyrosine (Y-374), are present at the same position in CYP3A163 suggesting the capability to oxidise larger or multiple substrates with likely differences towards substrate specificity. The signature haem binding motif of ten amino acids for all CYPs FXXGXXXCXG (FGAGPRNCLG, 442-451) with a basic amino acid (R) at the sixth position important for interaction with redox partners and the cysteine at eighth position is absolutely conserved in this decapeptide [34]. C-449 coordinates to the haem iron as a fifth ligand, necessary for the functional properties of CYPs to oxidize inactivated C-C or C–H bonds and electron transfer interactions [38], [39]. Beyond this conserved motif is another conserved tetrapeptide LQNF (467-470) [27].