Characterization of thiol-, aspartyl-, and thiol-metallo-peptidase activities in Madin-Darby canine kidney cells

Oliveira, Vitor; Ferro, Emer S.; Gomes, Marcelo D.; Oshiro, Maria E.M.; Almeida, Paulo C.; Juliano, María A.; Juliano, Luiz

doi:10.1002/(sici)1097-4644(20000301)76:3<478::aid-jcb14>3.3.co;2-8

Cited by 7 publications

(7 citation statements)

References 38 publications

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“…It is widely distributed in mammalian tissues with the highest expression levels in the brain, pituitary gland, and testis (5)(6)(7)(8). TOP is present in different subcellular locations depending on cell type, with reports of secreted and cytosolic forms (5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16), membrane association (5-7, 17, 18), and nuclear localization (7,12,14). Consistent with its broad tissue and subcellular compartment distribution, TOP appears to play a variety of physiological roles.…”

supporting

confidence: 69%

Crystal Structure of Human Thimet Oligopeptidase Provides Insight into Substrate Recognition, Regulation, and Localization

Ray¹,

Hines²,

Coll-Rodriguez

et al. 2004

Journal of Biological Chemistry

108

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Thimet oligopeptidase (TOP) is a zinc metallopeptidase that metabolizes a number of bioactive peptides and degrades peptides released by the proteasome, limiting antigenic presentation by MHC class I molecules. We present the crystal structure of human TOP at 2.0-Å resolution. The active site is located at the base of a deep channel that runs the length of the elongated molecule, an overall fold first seen in the closely related metallopeptidase neurolysin. Comparison of the two related structures indicates hinge-like flexibility and identifies elements near one end of the channel that adopt different conformations. Relatively few of the sequence differences between TOP and neurolysin map to the proposed substrate-binding site, and four of these variable residues may account for differences in substrate specificity. In addition, a loop segment (residues 599 -611) in TOP differs in conformation and degree of order from the corresponding neurolysin loop, suggesting it may also play a role in activity differences. Cysteines thought to mediate covalent oligomerization of rat TOP, which can inactivate the enzyme, are found to be surface-accessible in the human enzyme, and additional cysteines (residues 321,350, and 644) may also mediate multimerization in the human homolog. Disorder in the N terminus of TOP indicates it may be involved in subcellular localization, but a potential nuclear import element is found to be part of a helix and, therefore, unlikely to be involved in transport. A large acidic patch on the surface could potentially mediate a protein-protein interaction, possibly through formation of a covalent linkage.Thimet oligopeptidase (TOP, 1 3.4.24.15) is a 77-kDa zinc metalloendopeptidase that bears the His-Glu-Xaa-Xaa-His (HEXXH) active site sequence motif characteristic of a large superfamily of metallopeptidases (1-4). It is widely distributed in mammalian tissues with the highest expression levels in the brain, pituitary gland, and testis (5-8). TOP is present in different subcellular locations depending on cell type, with reports of secreted and cytosolic forms (5-16), membrane association (5-7, 17, 18), and nuclear localization (7,12,14). Consistent with its broad tissue and subcellular compartment distribution, TOP appears to play a variety of physiological roles. It has been implicated in the metabolism of a number of small peptides active in the central nervous system and the periphery including neurotensin, bradykinin, somatostatin, opioids, and angiotensin I (4, 6, 9, 16, 19 -26). In addition, recent reports demonstrate that TOP is primarily responsible for degrading peptides released from proteasomes, thereby limiting the extent of antigen presentation by MHC class I molecules (27-29). TOP has also been linked to amyloid precursor protein processing (30), and it promotes increased degradation of the A␤ peptide, a key component of amyloid plaques in Alzheimer's disease (31). Expression of TOP activity is regulated at the level of transcription (32-34), but activity may also be regulated by po...

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supporting

confidence: 69%

Crystal Structure of Human Thimet Oligopeptidase Provides Insight into Substrate Recognition, Regulation, and Localization

Ray¹,

Hines²,

Coll-Rodriguez

et al. 2004

Journal of Biological Chemistry

108

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“…Although the bulk of renal kininase activity can be attributed to NEP (138), a role for EP24.15 has also been postulated. Indeed, EP24.15 activity has recently been demonstrated in Madin-Darby canine kid-ney cells (139). In 1994, Yang et al (140) reported that cFP-AAY-pAB administration to rats decreases mean arterial pressure, while increasing renal blood flow, glomerular filtration rate, urine volume, and urinary sodium and potassium excretion.…”

Section: Cardiovascular/renal Homeostasismentioning

confidence: 99%

Soluble Metalloendopeptidases and Neuroendocrine Signaling

2002

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Peptidases play a vital and often highly specific role in the physiological and pathological generation and termination of peptide hormone signals. The thermolysin-like family of metalloendopeptidases involved in the extracellular processing of neuroendocrine and cardiovascular peptides are of particular significance, reflecting both their specificity for particular peptide substrates and their utility as therapeutic targets. Although the functions of the membrane-bound members of this family, such as angiotensin-converting enzyme and neutral endopeptidase, are well established, a role for the predominantly soluble family members in peptide metabolism is only just emerging. This review will focus on the biochemistry, cell biology, and physiology of the soluble metalloendopeptidases EC 3.4.24.15 (thimet oligopeptidase) and EC 3.4.24.16 (neurolysin), as well as presenting evidence that both peptidases play an important role in such diverse functions as reproduction, nociception, and cardiovascular homeostasis.

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“…We have previously reported that glioma C6 cells release an ep24.15-like activity into the culture medium (Ferro et al 1993). Since then, ep24.15 has been shown to be secreted in vivo from the median eminence (Lew et al 1997;Wu et al 1997) and by various cell lines Yamin et al 1999;Oliveira et al 2000). In agreement with the finding that ep24.15 is synthesized without a secretory signal peptide (Pierotti et al 1990), this enzyme is not present in secretory vesicles containing b-endorphin from pituitary AtT20 cells, from which its secretion can be stimulated by A23187 and corticotrophin-releasing hormone and blocked by brefeldin A and nocodazole .…”

mentioning

confidence: 99%

14‐3‐3 epsilon modulates the stimulated secretion of endopeptidase 24.15

Carreño

Goñi

Castro

et al. 2005

Journal of Neurochemistry

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Endopeptidase 24.15 (ep24.15: EC3.4.24.15), a secreted protein involved in peptide metabolism, is unusual in that it does not contain a signal peptide sequence. In this work, we describe the physical interaction between ep24.15 and 14-3-3 epsilon, one isoform of a family of ubiquitous phosphoserine/threoninescaffold proteins that organizes cell signaling and is involved in exocytosis. The interaction between ep24.15 and 14-3-3 epsilon increased following phosphorylation of ep24.15 at Ser 644 by protein kinase A (PKA). The co-localization of ep24.15 and 14-3-3 epsilon was increased by exposure of HEK293 cells (human embryonic kidney cells) to forskolin (10 lM). Overexpression of 14-3-3 epsilon in HEK293 cells almost doubled the secretion of ep24.15 stimulated by A23187 (7.5 lM) from 10% [1.4 ± 0.24 AFU/(min 10 6 cells)] to 19% [2.54 ± 0.24 AFU/(min 10 6 cells)] (p < 0.001) of the total intracellular enzyme activity. Treatment with forskolin had a synergistic effect on the A23187-stimulated secretion of ep24.15 that was totally blocked by the PKA inhibitor KT5720. The ep24.15 point mutation S644A reduced the co-localization of ep24.15 and 14-3-3 in stably transfected HEK293 cells. Indeed, secretion of the ep24.15 S644A mutant from these cells was only slightly stimulated by A23187 and insensitive to forskolin, in contrast to that of the wild type enzyme. Together, these data suggest that prior interaction with 14-3-3 is an important step in the unconventional stimulated secretion of ep24.15.

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Characterization of thiol-, aspartyl-, and thiol-metallo-peptidase activities in Madin-Darby canine kidney cells

Cited by 7 publications

References 38 publications

Crystal Structure of Human Thimet Oligopeptidase Provides Insight into Substrate Recognition, Regulation, and Localization

Crystal Structure of Human Thimet Oligopeptidase Provides Insight into Substrate Recognition, Regulation, and Localization

Soluble Metalloendopeptidases and Neuroendocrine Signaling

14‐3‐3 epsilon modulates the stimulated secretion of endopeptidase 24.15

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