Aspartate 496 from the subsite S2 drives specificity of human dipeptidyl peptidase III

Abramić, Marija; Karačić, Zrinka; Šemanjski, Maja; Vukelić, Bojana; Jajčanin-Jozić, Nina

doi:10.1515/hsz-2014-0247

Cited by 9 publications

(11 citation statements)

References 24 publications

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“…The bonds existence is confirmed by RDF analyses (right side of the Supplemental Figure S5A-C), where all the maximums are below 3 Å . This finding on important electrostatic interaction of Asp496 with aprotinin is in agreement with the recently reported results of mutational analysis which have shown that replacement of Asp496 with Gly lowered binding potency for aprotinin by 12.7-fold, compared to the wild-type hDPP III (Abrami c et al, 2015) indicating the important role of Asp496 in the hDPP III substrate specificity. -------GLU7 HI -------PRO8 HB, HI -------TYR10 HI -----HB -THR11 HI --HI HI HI HI…”

Section: Electrostatic Potential Surface and Solvent Accessible Surfasupporting

confidence: 92%

Aprotinin interacts with substrate-binding site of human dipeptidyl peptidase III

Agić

Brkić

Kazazić

et al. 2019

Journal of Biomolecular Structure and Dynamics

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Human dipeptidyl peptidase III (hDPP III) is a zinc-exopeptidase of the family M49 involved in final steps of intracellular protein degradation and in cytoprotective pathway Keap1-Nrf2. Biochemical and structural properties of this enzyme have been extensively investigated, but the knowledge on its contacts with other proteins is scarce. Previously, polypeptide aprotinin was shown to be a competitive inhibitor of hDPP III hydrolytic activity. In the present study, aprotinin was first investigated as a potential substrate of hDPP III, but no degradation products were demonstrated by MALDI-TOF mass spectrometry. Subsequently, molecular details of the protein-protein interaction between aprotinin and hDPP III were studied by molecular modeling. Docking and long molecular dynamics (MD) simulations have shown that aprotinin interacts by its canonical binding epitope with the substrate binding cleft of hDPP III. Thereby, free N-terminus of aprotinin is distant from the active-site zinc. Enzyme-inhibitor complex is stabilized by intermolecular hydrogen bonding network, electrostatic and hydrophobic interactions which mostly involve constituent amino acid residues of the hDPP III substrate binding subsites S1, S1', S2, S2' and S3'. This is the first study that gives insight into aprotinin binding to an metallopeptidase.

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Section: Electrostatic Potential Surface and Solvent Accessible Surfasupporting

confidence: 92%

Aprotinin interacts with substrate-binding site of human dipeptidyl peptidase III

Agić

Brkić

Kazazić

et al. 2019

Journal of Biomolecular Structure and Dynamics

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“…It has recently been shown that yeast DPP III does not prefer the canonical synthetic substrate of mammalian DPP III, Arg-Arg-arylamide [ 37 ]. In addition, it was reported that Asp496 situated in the S2 subsite of the human enzyme is an important determinant in the selectivity of human DPP III for Arg-Arg-2-naphthylamide (Arg 2 -2NA) [ 38 ]. The superposition of the crystal structures of Bt DPP III and hDPP III reveals that the residue that structurally corresponds to Asp496 is Asp465 in Bt DPP III.…”

Section: Resultsmentioning

confidence: 99%

Crystal structure of dipeptidyl peptidase III from the human gut symbiont Bacteroides thetaiotaomicron

et al. 2017

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Bacteroides thetaiotaomicron is a dominant member of the human intestinal microbiome. The genome of this anaerobe encodes more than 100 proteolytic enzymes, the majority of which have not been characterized. In the present study, we have produced and purified recombinant dipeptidyl peptidase III (DPP III) from B. thetaiotaomicron for the purposes of biochemical and structural investigations. DPP III is a cytosolic zinc-metallopeptidase of the M49 family, involved in protein metabolism. The biochemical results for B. thetaiotaomicron DPP III from our research showed both some similarities to, as well as certain differences from, previously characterised yeast and human DPP III. The 3D-structure of B. thetaiotaomicron DPP III was determined by X-ray crystallography and revealed a two-domain protein. The ligand-free structure (refined to 2.4 Å) was in the open conformation, while in the presence of the hydroxamate inhibitor Tyr-Phe-NHOH, the closed form (refined to 3.3 Å) was observed. Compared to the closed form, the two domains of the open form are rotated away from each other by about 28 degrees. A comparison of the crystal structure of B. thetaiotaomicron DPP III with that of the human and yeast enzymes revealed a similar overall fold. However, a significant difference with functional implications was discovered in the upper domain, farther away from the catalytic centre. In addition, our data indicate that large protein flexibility might be conserved in the M49 family.

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“…Although a significant knowledge on the molecular enzymology (structure-activity relationship) of the DPP III family accumulated in the last decade [19][20][21][22] , physiological roles of any of its members have not been elucidated in sufficient detail. This is partly due to the lack of specific inhibitors.…”

Section: Introductionmentioning

confidence: 99%

Novel dipeptidyl hydroxamic acids that inhibit human and bacterial dipeptidyl peptidase III

Cvitešić¹,

Sabljić

Makarević³

et al. 2016

Journal of Enzyme Inhibition and Medicinal Chemistry

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Human dipeptidyl peptidase III (hDPP III), a zinc-metallopeptidase of the family M49, is an activator of the Keap1-Nrf2 cytoprotective pathway involved in defense against oxidative stress. Pathophysiological roles of DPP III have not been elucidated yet, partly due to the lack of specific inhibitors. We showed that substrate analog H-Tyr-Phe-NHOH is a strong competitive inhibitor of hDPP III, while H-Tyr-Gly-NHOH expresses much weaker inhibition. To investigate the effects of amino acid substitutions in inhibitor P1 position, we synthesized three new dipeptidyl hydroxamates and examined their influence on the activity of hDPP III and DPP III from the human gut symbiont Bacteroides thetaiotaomicron. The extent of inhibition of hDPP III, but not of bacterial enzyme, was dependent on the amino acid in P1. H-Phe-Phe-NHOH is recognized as one of the strongest inhibitors of hDPP III (K i ¼ 0.028 mM), and H-Phe-Leu-NHOH discriminated between human and bacterial ortholog of the M49 family.

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Aspartate 496 from the subsite S2 drives specificity of human dipeptidyl peptidase III

Cited by 9 publications

References 24 publications

Aprotinin interacts with substrate-binding site of human dipeptidyl peptidase III

Aprotinin interacts with substrate-binding site of human dipeptidyl peptidase III

Crystal structure of dipeptidyl peptidase III from the human gut symbiont Bacteroides thetaiotaomicron

Novel dipeptidyl hydroxamic acids that inhibit human and bacterial dipeptidyl peptidase III

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