Crystal Structure and Mechanism of Tripeptidyl Activity of Prolyl Tripeptidyl Aminopeptidase from Porphyromonas gingivalis

Itō, Kiyoshi; Nakajima, Yoshitaka; Xu, Yue; Yamada, N.; Onohara, Yuko; Ito, Takashi; Matsubara, Futoshi; Kabashima, Tsutomu; Nakayama, Koji; Yoshimoto, Tadashi

doi:10.1016/j.jmb.2006.06.083

Cited by 27 publications

(33 citation statements)

References 41 publications

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“…The Cα positions of the three structures could be superimposed onto that of PTP39 6 with root-mean-square deviations of 0.16, 0.16, and 0.16 Å, and maximum distances of 1.92, 1.18, and 1.50 Å, respectively. Briefly, every subunit in the homodimer consists of two domains, the eight-bladed β-propeller domain (Pro51-Lys470) and the catalytic domain (Asn471-Leu732).…”

Section: Overall Structuresmentioning

confidence: 99%

“…One of the hydroxyl groups of borate formed a hydrogen bond with the hydroxyl group of Tyr518 at a distance of 2.65 Å. We previously reported the structure of the S603A mutant complexed with the substrate Gly-Ala-Pro-βNA (S603A-substrate complex; PDB code: 2dcm) 6 . From the comparison between these two structures, the position of the inhibitor was found closer to the protein than that of the substrate (Figure 3a).…”

Section: Active Site In Ptp39-inhibitor Complexmentioning

confidence: 99%

“…We previously determined the structures of PTP39 and its S603A mutant complexed with a substrate, Gly-Ala-Pro-βNA 6 . In that study, we suggested that the Pro residue of substrates is accommodated by a hydrophobic pocket composed of Tyr604, Val629, Trp632, Tyr635, Tyr639, Val680, and Val681, and that the N-terminal amino group is recognized by Glu205 and Glu636.…”

Section: The Inhibitor Binding To the Active Sitementioning

confidence: 99%

“…PTP shows no specificity for residues at the P2 position; for instance, it has been reported that PTP can hydrolyze substrates containing nonpolar aliphatic (Ala, Gly, Pro, and Val), aromatic (Phe), polar uncharged (Asn), and charged residues (Glu, Arg, and His) at the P2 position 4,6 . The large space of the S2 site would not limit the kind of residue at the P2 position of substrates.…”

Section: The Inhibitor Binding To the Active Sitementioning

confidence: 99%

“…Recently, we determined the crystal structures of PTP with an N-terminal truncation of 38 residues (PTP39) and its S603A mutant enzyme complexed with the substrate, Gly-Ala-Pro-β-naphthylamide (-βNA), at 2.1 and 2.9 Å resolution, Prolyl Tripeptidyl Aminopeptidase from P. gingivalis 5 respectively 6 . The subunit of the enzyme is composed of two domains, the N-terminal eight-bladed β-propeller domain (Glu45-Lys470) and the C-terminal catalytic domain (Asn471-Leu732).…”

Section: Introductionmentioning

confidence: 99%

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Novel Inhibitor for Prolyl Tripeptidyl Aminopeptidase from Porphyromonas gingivalis and Details of Substrate-recognition Mechanism

Xu¹,

Nakajima²,

Itō³

et al. 2008

Journal of Molecular Biology

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Prolyl Tripeptidyl Aminopeptidase from P. gingivalis 2 SummaryA new inhibitor, H-Ala-Ile-pyrrolidin-2-yl boronic acid, was developed as an inhibitor against prolyl tripeptidyl aminopeptidase with the K i value of 88.1 nM. The structure of the prolyl tripeptidyl aminopeptidase complexed with the inhibitor (enzyme-inhibitor complex) was determined at 2.2 Å resolution. The inhibitor was bound to the active site through a covalent bond between Ser603 and the boron atom of the inhibitor. This structure should closely mimic the structure of the reaction-intermediate between the enzyme and substrate.We previously proposed that two glutamate residues, Glu205 and Glu636, are involved in the recognition of substrates. In order to clarify the function of these glutamate residues in substrate recognition, three mutant enzymes, E205A, E205Q, and E636A were generated by site-directed mutagenesis. The E205A mutant was expressed as an inclusion body. The E205Q mutant was expressed in soluble form, but no activity was detected for it.Here, the structures of the E636A mutant and its complex with the inhibitor were determined. The inhibitor was located at almost the same position as in the wild-type enzyme-inhibitor complex. The amino group of the inhibitor interacted with Glu205 and the main-chain carbonyl group of Gln203. In addition, a water molecule in the place of Glu636 of the wild-type enzyme interacted with the amino group of the inhibitor. This water molecule was located near the position of Glu636 in the wild type and formed a hydrogen bond with Gln203. The k cat /K M values of the E636A mutant toward the two substrates used were smaller than those of the wild type by two Prolyl Tripeptidyl Aminopeptidase from P. gingivalis 3 orders of magnitude. The K i value of our inhibitor for the E636A mutant was 48.8 M, which was 554-fold higher than that against the wild-type enzyme. Consequently, it was concluded that Glu205 and Glu636 are significant residues for the N-terminal recognition of a substrate.

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Section: Overall Structuresmentioning

confidence: 99%

Section: Active Site In Ptp39-inhibitor Complexmentioning

confidence: 99%

Section: The Inhibitor Binding To the Active Sitementioning

confidence: 99%

Section: The Inhibitor Binding To the Active Sitementioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Novel Inhibitor for Prolyl Tripeptidyl Aminopeptidase from Porphyromonas gingivalis and Details of Substrate-recognition Mechanism

Xu¹,

Nakajima²,

Itō³

et al. 2008

Journal of Molecular Biology

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Structural evidence that puromycin hydrolase is a new type of aminopeptidase with a prolyl oligopeptidase family fold

et al. 2011

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Computational analysis of the domain architecture and substrate-gating mechanism of prolyl oligopeptidases from Shewanella woodyi and identification probable lead molecules

Patil

Skariyachan

Mutt

et al. 2015

Interdiscip Sci Comput Life Sci

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Prolyl oligopeptidases (POP) are serine proteases found in prokaryotes and eukaryotes which hydrolyze the peptide bond containing proline. The current study focuses on the analysis of POP sequences, their distribution and domain architecture in Shewanella woodyi, a Gram negative, luminous bacterium which causes celiac sprue and similar infections in marine organisms. The POP undergoes huge inter-domain movement, which allows possible route for the entry of any substrate. Hence, it offers an opportunity to understand the mechanism of substrate gating by studying the domain architecture and possibility to identify a probable drug target. In the present study, the POP sequence was retrieved from GenBank data base and the best homologous templates were identified by PSI-BLAST search. The three dimensional structures of the closed and open forms of POP from Shewanella woodyi, which are not available in native form, was generated by homology modeling. The ideal lead molecules were screened by computer aided virtual screening and the binding potential of the best leads towards the target was studied by molecular docking. The domain architecture of the POP revealed that, it has a propeller domain consist of β-sheets, surrounded by α-helices and α/β hydrolase domain with catalytic triad containing Ser-564, Asp-646 and His-681. The hypothetical models of open and closed POP showed backbone RMSD value of 0.56 Å and 0.65 Å respectively. Ramachandran plot of the open and closed POP conformations accounts for 99.4% and 98.7% residues in the favoured region respectively. Our study revealed that, propeller domain comes as an insert between N-terminal and C-terminal α/β hydrolase domain. Molecular docking, drug likeliness properties and ADME prediction suggested that KUC-103481N and Pramiracetum can be used as probable lead molecules towards the POP from Shewanella woodyi.

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Crystal Structure and Mechanism of Tripeptidyl Activity of Prolyl Tripeptidyl Aminopeptidase from Porphyromonas gingivalis

Cited by 27 publications

References 41 publications

Novel Inhibitor for Prolyl Tripeptidyl Aminopeptidase from Porphyromonas gingivalis and Details of Substrate-recognition Mechanism

Novel Inhibitor for Prolyl Tripeptidyl Aminopeptidase from Porphyromonas gingivalis and Details of Substrate-recognition Mechanism

Structural evidence that puromycin hydrolase is a new type of aminopeptidase with a prolyl oligopeptidase family fold

Computational analysis of the domain architecture and substrate-gating mechanism of prolyl oligopeptidases from Shewanella woodyi and identification probable lead molecules

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