The structure and function of human dipeptidyl peptidase IV, possessing a unique eight-bladed β-propeller fold

Hiramatsu, Hidenori; Kyono, Kiyoshi; Higashiyama, Yutaka; Fukushima, Chiaki; Shima, Hideaki; Sugiyama, Shigeru; Inaka, Koji; Yamamoto, Atsushi; Shimizu, Ryo

doi:10.1016/s0006-291x(03)00258-4

Cited by 89 publications

(57 citation statements)

References 21 publications

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“…Most interesting, coordination of a water molecule (Wat 123 , see Fig. 4A) by the tyrosine was observed in different apo structures (previously published human DPP-IV (Protein Data Bank code 1PFQ (24) and Protein Data Bank code 1J2E (25)) and porcine DPP-IV (Protein Data Bank code 1ORV (26))) as well as our own in-house information) but not in the phenylalanine mutant. In one of the previously published human apo structures (Protein Data Bank code 1NU6 (27)), this water molecule is absent or not included.…”

Section: Structural and Sequence Analysis Of The Active Site Of Dpp-iv-mentioning

confidence: 56%

Tyrosine 547 Constitutes an Essential Part of the Catalytic Mechanism of Dipeptidyl Peptidase IV

Bjelke

Christensen

Branner³

et al. 2004

Journal of Biological Chemistry

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Human dipeptidyl peptidase IV (DPP-IV) is a ubiquitously expressed type II transmembrane serine protease. It cleaves the penultimate positioned prolyl bonds at the N terminus of physiologically important peptides such as the incretin hormones glucagon-like peptide 1 and glucose-dependent insulinotropic peptide. In this study, we have characterized different active site mutants. The Y547F mutant as well as the catalytic triad mutants S630A, D708A, and H740L showed less than 1% wild type activity. X-ray crystal structure analysis of the Y547F mutant revealed no overall changes compared with wild type apoDPP-IV, except the ablation of the hydroxyl group of Tyr 547 and a water molecule positioned in close proximity to Tyr 547 . To elucidate further the reaction mechanism, we determined the crystal structure of DPP-IV in complex with diisopropyl fluorophosphate, mimicking the tetrahedral intermediate. The kinetic and structural findings of the tyrosine residue are discussed in relation to the catalytic mechanism of DPP-IV and to the inhibitory mechanism of the 2-cyanopyrrolidine class of potent DPP-IV inhibitors, proposing an explanation for the specificity of this class of inhibitors for the S9b family among serine proteases.

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Section: Structural and Sequence Analysis Of The Active Site Of Dpp-iv-mentioning

confidence: 56%

Tyrosine 547 Constitutes an Essential Part of the Catalytic Mechanism of Dipeptidyl Peptidase IV

Bjelke

Christensen

Branner³

et al. 2004

Journal of Biological Chemistry

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“…The N-terminal free amino group of the substrate is recognized by two glutamate residues, Glu205* and Glu206* (the * denotes the residue from DPIV) at the β-propeller domain [9][10][11][12][17][18] . Glu205* and Glu206* are found in the helix appended to the fourth blade.…”

Section: Function Of the Glutamate Residues At The Active Sitementioning

confidence: 99%

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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“…It is generally known that the amide nitrogen of the main chain plays a role as an oxyanion hole. In addition, it has been reported that prolyl oligopeptidase and dipeptidyl peptidase IV, both of which belong to family S9, possess a tyrosine residue that acts as the oxyanion hole (31)(32)(33)(34). It is likely that the tyrosine residues in family M1 exert a function similar to forming an oxyanion hole in serine proteases.…”

mentioning

confidence: 99%

Crystal Structure of Aminopeptidase N (Proteobacteria Alanyl Aminopeptidase) from Escherichia coli and Conformational Change of Methionine 260 Involved in Substrate Recognition

Itō¹,

Nakajima²,

Onohara

et al. 2006

Journal of Biological Chemistry

132

151

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The structure and function of human dipeptidyl peptidase IV, possessing a unique eight-bladed β-propeller fold

Cited by 89 publications

References 21 publications

Tyrosine 547 Constitutes an Essential Part of the Catalytic Mechanism of Dipeptidyl Peptidase IV

Tyrosine 547 Constitutes an Essential Part of the Catalytic Mechanism of Dipeptidyl Peptidase IV

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

Crystal Structure of Aminopeptidase N (Proteobacteria Alanyl Aminopeptidase) from Escherichia coli and Conformational Change of Methionine 260 Involved in Substrate Recognition

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