Molecular determinants of human dipeptidyl peptidase III sensitivity to thiol modifying reagents
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Zrinka Karačić
Abstract
Human dipeptidyl peptidase III (DPP III) is a member of the metallopeptidase family M49, involved in protein metabolism and oxidative stress response. DPP III crystal structure shows the two lobe-like domains separated by a wide cleft. The human enzyme has a total of six cysteines, three in the lower (Cys19, Cys147, and Cys176) and three in the upper (Cys509, Cys519, and Cys654), catalytic, domain containing the active-site zinc ion. To elucidate the molecular basis of this enzyme’s susceptibility to sulfhydryl reagents, biochemical analysis of a set of Cys to Ala mutants was used, supported by mass spectrometry. Cys176, a residue 44 Å apart from the catalytic center of the ligand-free enzyme, was found responsible for the inactivation with the submicromolar concentration of an organomercurial compound, and three additional cysteines contri\xadbuted to sensitivity to aromatic disulfides. Upon treatment with oxidized glutathione [glutathione disulfide (GSSG)], cysteine residues at positions 147, 176, and 654 were found glutathionylated. The mutational analysis confirmed the involvement of Cys176 and Cys654 in human DPP III inactivation by GSSG. Observation that Cys176, a residue quite distant from the active center, contributes to enzyme inactivation, indicates that the substrate-binding site of human DPP III comprises both lower and upper protein domain.
©2012 by Walter de Gruyter Berlin Boston
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Artikel in diesem Heft
- Masthead
- Masthead
- Guest Editorial
- Highlight: 29th Winter School on Proteinases and Their Inhibitors
- Highlight: 29th Winter School on Proteinases and their Inhibitors
- Genetic polymorphisms in the human tissue kallikrein (KLK) locus and their implication in various malignant and non-malignant diseases
- Effects of cathepsin K deficiency on intercellular junction proteins, luminal mucus layers, and extracellular matrix constituents in the mouse colon
- Three-dimensional cultures modeling premalignant progression of human breast epithelial cells: role of cysteine cathepsins
- Cysteine cathepsins are not critical for TRAIL- and CD95-induced apoptosis in several human cancer cell lines
- Dissecting the impact of Frizzled receptors in Wnt/β-catenin signaling of human mesenchymal stem cells
- Overexpression of the urokinase receptor splice variant uPAR-del4/5 in breast cancer cells affects cell adhesion and invasion in a dose-dependent manner and modulates transcription of tumor-associated genes
- IGF-I receptor phosphorylation is impaired in cathepsin X-deficient prostate cancer cells
- Cell surface engineering of renal cell carcinoma with glycosylphosphatidylinositol-anchored TIMP-1 blocks TGF-β1 activation and reduces regulatory ID gene expression
- Gingipain aminopeptidase activities in Porphyromonas gingivalis
- CLIPPER: an add-on to the Trans-Proteomic Pipeline for the automated analysis of TAILS N-terminomics data
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- Characterization of angiotensin I-converting enzyme N-domain selectivity using positional-scanning combinatorial libraries of fluorescence resonance energy transfer peptides
- Mast cells limit extracellular levels of IL-13 via a serglycin proteoglycan-serine protease axis
