Dipeptidyl Aminopeptidase IV fromStenotrophomonas maltophiliaExhibits Activity against a Substrate Containing a 4-Hydroxyproline Residue

Abstract

The crystal structure of dipeptidyl aminopeptidase IV from Stenotrophomonas maltophilia was determined at 2.8-Å resolution by the multiple isomorphous replacement method, using platinum and selenomethionine derivatives. The crystals belong to space group P4 3 2 1 2, with unit cell parameters a ‫؍‬ b ‫؍‬ 105.9 Å and c ‫؍‬ 161.9 Å. Dipeptidyl aminopeptidase IV is a homodimer, and the subunit structure is composed of two domains, namely, N-terminal ␤-propeller and C-terminal catalytic domains. At the active site, a hydrophobic pocket to accommodate a proline residue of the substrate is conserved as well as those of mammalian enzymes. Stenotrophomonas dipeptidyl aminopeptidase IV exhibited activity toward a substrate containing a 4-hydroxyproline residue at the second position from the N terminus. In the Stenotrophomonas enzyme, one of the residues composing the hydrophobic pocket at the active site is changed to Asn611 from the corresponding residue of Tyr631 in the porcine enzyme, which showed very low activity against the substrate containing 4-hydroxyproline. The N611Y mutant enzyme was generated by site-directed mutagenesis. The activity of this mutant enzyme toward a substrate containing 4-hydroxyproline decreased to 30.6% of that of the wild-type enzyme. Accordingly, it was considered that Asn611 would be one of the major factors involved in the recognition of substrates containing 4-hydroxyproline.

Structural Insights into the Pseudomonas aeruginosa Type VI Virulence Effector Tse1 Bacteriolysis and Self-protection Mechanisms

“…4C). DALI searches (23) with the structure of Tsi1 as the bait obtained several structures containing a typical ␤-propeller fold, such as dipeptidyl aminopeptidase IV from Stenotrophomonas maltophilia (Z ϭ 7.2, PDB code 2ECF) (30). Although the structure of Tsi1 can be roughly superimposed with part of the characterized ␤-propeller fold (supplemental Fig.…”

Activity and selectivity cliffs for DPP‐IV inhibitors: Lessons we can learn from SAR studies and their application to virtual screening

“…Stenotrophomonas maltophilia as a template (PDB code 2ECF) 134 and is available from the ModBase database 135 using access numbers 37577089 136 and 123983020 137 for DPP8 and DPP9, respectively. The second set of homology models was reported by Janardhan and Reddy 132 and built using the A chain of a human DPP-IV structure (PDB code 1 × 70) as a template.…”

Periplasmic form of dipeptidyl aminopeptidase IV fromPseudoxanthomonas mexicanaWO24: purification, kinetic characterization, crystallization and X-ray crystallographic analysis

“…Initial phase determination for the PmDAP IV crystal was performed by the molecular-replacement technique using the coordinates of one protomer of SmDAP IV (PDB entry 2ecf; Nakajima et al, 2008), which has approximately 74% aminoacid sequence identity to PmDAP IV, as a search model. Bound water molecules were removed from the search model.…”

“…To date, several crystal structures of mammalian DPP IVs have been reported and substrate-recognition mechanisms of the mammalian DPP IVs have been discussed in detail (Engel et al, 2003;Hiramatsu et al, 2003;Rasmussen et al, 2003). However, crystal structure analysis of a bacterial DAP IV has been only reported at medium resolution (2.8 Å ) for DAP IV from S. maltophilia (SmDAP IV; Nakajima et al, 2008) in a peptide-free form. The structural analysis of SmDAP IV revealed that the overall structure of SmDAP IV is similar to those of mammalian DPP IVs; however, an activesite arginine residue (Arg125 in human DPP IV) which is responsible for the recognition of the carbonyl group of the P2 residue of a substrate peptide is not conserved in bacterial DAP IVs.…”