Structural definition and substrate specificity of the S28 protease family: the crystal structure of human prolylcarboxypeptidase

Soisson, S.M.; Patel, Sangita; Abeywickrema, Pravien; Byrne, Noel; Diehl, Ronald E.; Hall, Dawn L.; Ford, Rachael E.; Reid, John C.; Rickert, Keith; Shipman, Jennifer M.; Sharma, Sujata; Lumb, Kevin J.

doi:10.1186/1472-6807-10-16

Cited by 50 publications

(51 citation statements)

References 37 publications

(38 reference statements)

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“…In DPP IV and DPP VII, for example, subsite S1 is small and hydrophobic, restricting P1 to proline, alanine, or glycine (38). In contrast, all subsites in DPP III are deep and somewhat hydrophobic (SI Appendix, Fig.…”

Section: Resultsmentioning

confidence: 99%

Entropy-driven binding of opioid peptides induces a large domain motion in human dipeptidyl peptidase III

Bezerra

Dobrovetsky

Viertlmayr

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

161

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Opioid peptides are involved in various essential physiological processes, most notably nociception. Dipeptidyl peptidase III (DPP III) is one of the most important enkephalin-degrading enzymes associated with the mammalian pain modulatory system. Here we describe the X-ray structures of human DPP III and its complex with the opioid peptide tynorphin, which rationalize the enzyme's substrate specificity and reveal an exceptionally large domain motion upon ligand binding. Microcalorimetric analyses point at an entropy-dominated process, with the release of water molecules from the binding cleft ("entropy reservoir") as the major thermodynamic driving force. Our results provide the basis for the design of specific inhibitors that enable the elucidation of the exact role of DPP III and the exploration of its potential as a target of pain intervention strategies.isothermal titration calorimetry | metallopeptidase | peptide binding | X-ray crystallography T he endogenous opioid system, composed of opioid peptides and their receptors, modulates a large number of physiological processes, such as endocrine and immune function, gastrointestinal motility, respiration, reward, stress, complex social behavior (e.g., sexual activity), vulnerability to drug addiction, and most notably the procession and transmission of pain stimuli (nociception) (1, 2). Two major types of endogenous opioid peptides are those containing enkephalin sequences at the N terminus (TyrGly-Gly-Phe-Met/Leu) (3) and, more recently identified, endomorphins 1 and 2 (Tyr-Pro-Trp/Phe-Phe-NH 2 ) (4, 5). Knowledge and control over synthesis and degradation pathways of this class of molecules is prerequisite for the development of new therapies that target pertinent physiological processes.Dipeptidyl peptidase III (DPP III), also known as enkephalinase B, is an enkephalin-degrading enzyme that cleaves dipeptides sequentially from the N termini of substrates (6). All DPP IIIs described thus far contain the unique zinc-binding motif HEXXGH characteristic of metallopeptidase family M49 (7). Enzymes from several human and animal tissues, as well as from lower eukaryotes, were purified and biochemically characterized (8, 9). DPP III is largely found as a cytosolic protein, although membrane association in rat brain and Drosophila melanogaster has been described (10, 11). The 3D structure of the yeast ortholog has recently been determined, revealing a unique protein fold with two lobes forming a wide-open substrate-binding cleft (12). The lack of structural information on peptide complexes, however, left the question of substrate specificity largely unanswered.DPP III purified from monkey brain microsomes is strongly inhibited by the neuropeptide spinorphin (Leu-Val-Val-Tyr-ProTrp-Thr), an endogenous factor isolated from bovine spinal cord that also inhibits other enkephalin-degrading enzymes, such as neutral endopeptidase (NEP, neprilysin), aminopeptidase, and angiotensin-converting enzyme (13). Because of a different mode of action compared with morphine, spinorp...

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Section: Resultsmentioning

confidence: 99%

Entropy-driven binding of opioid peptides induces a large domain motion in human dipeptidyl peptidase III

Bezerra

Dobrovetsky

Viertlmayr

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

161

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“…In normal animals, PRCP contributes to the production of bradykinin and angiotensin. [1][2][3][4][5][6][7] Both peptides bind to their constitutive receptors, the B2R and Mas, to contribute to the constitutive level of vascular NO that contributes to thrombosis protection ( Figure 7). Alternatively, PRCP gt/gt mice are hypertensive.…”

Section: Discussionmentioning

confidence: 99%

“…6 The molecular affinity of PRCP for Pro-X bonds is demonstrated by its crystal structure with the identification of hydrophobic residues near the substrate proline at the active site. 1,2 PRCP is up-regulated during the angiogenic processes of vascular development. 7 The exact physiologic role for PRCP is not completely known.…”

Section: Introductionmentioning

confidence: 99%

“…10 PRCP is linked to metabolic syndrome in humans. 11 PRCP also degrades ␣-melanocyte stimulating hormone (␣-MSH [1][2][3][4][5][6][7][8][9][10][11][12][13] ) to its inactive ␣-MSH [1][2][3][4][5][6][7][8][9][10][11][12] form by cleaving its C-terminal Val. 12 ␣-MSH [1][2][3][4][5][6][7][8][9][10][11][12][13] stimulates an anorexigenic response.…”

Section: Introductionmentioning

confidence: 99%

“…PRCP dimerizes and has a unique protease structure with closest identity to dipeptidyl peptidase 7 with a novel helical structural domain (SKS domain) that caps the active site of the catalytic Asp-His-Ser triad. 1,2 The protein was first isolated from swine kidney lysosomal fractions and identified as a bradykinin and angiotensin II carboxypeptidase. 3 PRCP is found on the surface of endothelial cells where it is a high-affinity activator that converts prekallikrein bound to high molecular weight kininogen to the serine protease plasma kallikrein.…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Murine prolylcarboxypeptidase depletion induces vascular dysfunction with hypertension and faster arterial thrombosis

Adams¹,

LaRusch²,

Stavrou³

et al. 2011

Blood

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Structural and time‐resolved mechanistic investigations of protein hydrolysis by the acidic proline‐specific endoprotease from Aspergillus niger

Pijning,

Vujičić‐Žagar,

van der Laan

et al. 2023

Protein Science

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Proline‐specific endoproteases have been successfully used in, for example, the in‐situ degradation of gluten, the hydrolysis of bitter peptides, the reduction of haze during beer production, and the generation of peptides for mass spectroscopy and proteomics applications. Here we present the crystal structure of the extracellular proline‐specific endoprotease from Aspergillus niger (AnPEP), a member of the S28 peptidase family with rarely observed true proline‐specific endoprotease activity. Family S28 proteases have a conventional Ser‐Asp‐His catalytic triad, but their oxyanion‐stabilizing hole shows a glutamic acid, an amino acid not previously observed in this role. Since these enzymes have an acidic pH optimum, the presence of a glutamic acid in the oxyanion hole may confine their activity to an acidic pH. Yet, considering the presence of the conventional catalytic triad, it is remarkable that the A. niger enzyme remains active down to pH 1.5. The determination of the primary cleavage site of cytochrome c along with molecular dynamics‐assisted docking studies indicate that the active site pocket of AnPEP can accommodate a reverse turn of approximately 12 amino acids with proline at the S1 specificity pocket. Comparison with the structures of two S28‐proline‐specific exopeptidases reveals not only a more spacious active site cavity but also the absence of any putative binding sites for amino‐ and carboxyl‐terminal residues as observed in the exopeptidases, explaining AnPEP's observed endoprotease activity.

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Structural definition and substrate specificity of the S28 protease family: the crystal structure of human prolylcarboxypeptidase

Cited by 50 publications

References 37 publications

Entropy-driven binding of opioid peptides induces a large domain motion in human dipeptidyl peptidase III

Entropy-driven binding of opioid peptides induces a large domain motion in human dipeptidyl peptidase III

Murine prolylcarboxypeptidase depletion induces vascular dysfunction with hypertension and faster arterial thrombosis

Structural and time‐resolved mechanistic investigations of protein hydrolysis by the acidic proline‐specific endoprotease from Aspergillus niger

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