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Nature Communications Feb 2022The proteasome recognizes ubiquitinated proteins and can also edit ubiquitin marks, allowing substrates to be rejected based on ubiquitin chain topology. In yeast,...
The proteasome recognizes ubiquitinated proteins and can also edit ubiquitin marks, allowing substrates to be rejected based on ubiquitin chain topology. In yeast, editing is mediated by deubiquitinating enzyme Ubp6. The proteasome activates Ubp6, whereas Ubp6 inhibits the proteasome through deubiquitination and a noncatalytic effect. Here, we report cryo-EM structures of the proteasome bound to Ubp6, based on which we identify mutants in Ubp6 and proteasome subunit Rpt1 that abrogate Ubp6 activation. The Ubp6 mutations define a conserved region that we term the ILR element. The ILR is found within the BL1 loop, which obstructs the catalytic groove in free Ubp6. Rpt1-ILR interaction opens the groove by rearranging not only BL1 but also a previously undescribed network of three interconnected active-site-blocking loops. Ubp6 activation and noncatalytic proteasome inhibition are linked in that they are eliminated by the same mutations. Ubp6 and ubiquitin together drive proteasomes into a unique conformation associated with proteasome inhibition. Thus, a multicomponent allosteric switch exerts simultaneous control over both Ubp6 and the proteasome.
Topics: Adenosine Triphosphatases; Catalytic Domain; Cryoelectron Microscopy; Cytoplasm; Endopeptidases; Proteasome Endopeptidase Complex; Protein Conformation; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Ubiquitin; Ubiquitinated Proteins
PubMed: 35149681
DOI: 10.1038/s41467-022-28186-y -
Biochemistry Sep 2022Cathepsin B is a lysosomal protease that participates in protein degradation. However, cathepsin B is also active under neutral pH conditions of the cytosol, nuclei, and...
Cathepsin B is a lysosomal protease that participates in protein degradation. However, cathepsin B is also active under neutral pH conditions of the cytosol, nuclei, and extracellular locations. The dipeptidyl carboxypeptidase (DPCP) activity of cathepsin B, assayed with the Abz-GIVR↓AK(Dnp)-OH substrate, has been reported to display an acidic pH optimum. In contrast, the endopeptidase activity, monitored with Z-RR-↓AMC, has a neutral pH optimum. These observations raise the question of whether other substrates can demonstrate cathepsin B DPCP activity at neutral pH and endopeptidase activity at acidic pH. To address this question, global cleavage profiling of cathepsin B with a diverse peptide library was conducted under acidic and neutral pH conditions. Results revealed that cathepsin B has (1) major DPCP activity and modest endopeptidase activity under both acidic and neutral pH conditions and (2) distinct pH-dependent amino acid preferences adjacent to cleavage sites for both DPCP and endopeptidase activities. The pH-dependent cleavage preferences were utilized to design a new Abz-GVR↓AK(Dnp)-OH DPCP substrate, with norleucine (n) at the P3 position, having improved DPCP activity of cathepsin B at neutral pH compared to the original Abz-GIVR↓AK(Dnp)-OH substrate. The new Z-VR-AMC and Z-ER-AMC substrates displayed improved endopeptidase activity at acidic pH compared to the original Z-RR-AMC. These findings illustrate the new concept that cathepsin B possesses DPCP and endopeptidase activities at both acidic and neutral pH values. These results advance understanding of the pH-dependent cleavage properties of the dual DPCP and endopeptidase activities of cathepsin B that function under different cellular pH conditions.
Topics: Cathepsin B; Cathepsins; Endopeptidases; Hydrogen-Ion Concentration; Hydrolysis; Proteolysis; Substrate Specificity
PubMed: 35981509
DOI: 10.1021/acs.biochem.2c00358 -
Current Biology : CB
Review
Topics: Animals; Catalysis; Cysteine Endopeptidases; Humans; Multienzyme Complexes; Proteasome Endopeptidase Complex; Proteins; Ubiquitins
PubMed: 9889109
DOI: 10.1016/s0960-9822(98)00004-9 -
Translational Neurodegeneration 2020Mammalian asparagine endopeptidase (AEP) is a cysteine protease that cleaves its protein substrates on the C-terminal side of asparagine residues. Converging lines of... (Review)
Review
Mammalian asparagine endopeptidase (AEP) is a cysteine protease that cleaves its protein substrates on the C-terminal side of asparagine residues. Converging lines of evidence indicate that AEP may be involved in the pathogenesis of several neurological diseases, including Alzheimer's disease, Parkinson's disease, and frontotemporal dementia. AEP is activated in the aging brain, cleaves amyloid precursor protein (APP) and promotes the production of amyloid-β (Aβ). We renamed AEP to δ-secretase to emphasize its role in APP fragmentation and Aβ production. AEP also cleaves other substrates, such as tau, α-synuclein, SET, and TAR DNA-binding protein 43, generating neurotoxic fragments and disturbing their physiological functions. The activity of δ-secretase is tightly regulated at both the transcriptional and posttranslational levels. Here, we review the recent advances in the role of δ-secretase in neurodegenerative diseases, with a focus on its biochemical properties and the transcriptional and posttranslational regulation of its activity, and discuss the clinical implications of δ-secretase as a diagnostic biomarker and therapeutic target for neurodegenerative diseases.
Topics: Amyloid Precursor Protein Secretases; Amyloid beta-Peptides; Animals; Cysteine Endopeptidases; Humans; Neurodegenerative Diseases
PubMed: 31911834
DOI: 10.1186/s40035-019-0179-3 -
Journal of Biochemistry Feb 1998The proteasome is a multisubunit protease complex with an apparent sedimentation coefficient of 20S. Two types of regulatory complexes, named PA700 and PA28, bind to... (Review)
Review
The proteasome is a multisubunit protease complex with an apparent sedimentation coefficient of 20S. Two types of regulatory complexes, named PA700 and PA28, bind to both ends of the cylindrical 20S proteasome to form the dumbbell-like and football-like proteasomes, respectively. The former complex, named the 26S proteasome, is a eukaryotic ATP-dependent protease and appears to be well organized as a large complex of 2 MDa, consisting of approximately 40 polypeptides, to facilitate rapid proteolysis. It is assumed to be a protein "death machine", destroying a variety of cellular proteins that have acquired a specific degradation signal(s) such as a multiubiquitin chain. Recently data on in vivo substrates for the ubiquitin-proteasome pathway have been accumulating rapidly, implying its involvement in many biologically important processes, such as cell-cycle regulation, signal transduction, protein quality control, and the immune response. The newly-identified PA28 family proteins are inducible by interferons, and may cooperate with the 26S proteasome or play additional roles. Since the proteasome is capable of catalyzing breakdown of proteins not only irreversibly, but also rapidly and timely, it is thought to be a new regulatory system for biological reactions in eukaryotes.
Topics: Animals; Cysteine Endopeptidases; Humans; Multienzyme Complexes; Proteasome Endopeptidase Complex; Structure-Activity Relationship
PubMed: 9538192
DOI: 10.1093/oxfordjournals.jbchem.a021922 -
Expert Opinion on Therapeutic Targets Oct 2017Fibroblast activation protein-α (FAP-α) belongs to the family of prolyl-specific serine proteases. FAP-α displays both exopeptidase and... (Review)
Review
Fibroblast activation protein-α (FAP-α) belongs to the family of prolyl-specific serine proteases. FAP-α displays both exopeptidase and endopeptidase/gelatinase/collagenase activities. FAP-α protein and/or activity have been associated with fibrosis, inflammation and cancer, but the protein is undetectable in most normal tissues. FAP-α is selectively expressed at sites of tissue remodeling and repair and enhances tumor progression, suggesting that this protease may be a therapeutic target to treat human disorders associated with fibrotic dysregulation. Areas covered: In this review, we summarize the mechanisms driving tissue fibrosis and describe some of the enzymes involved in fibrosis, concentrating on FAP-α. We describe its enzymatic properties, discuss the tools developed to control its activity and the problem of selectivity toward the other proteases of the family and outline its potential biological substrates. We also consider non-enzymatic functions of this protein and suggest that repression of FAP-α expression may represent therapeutic options. Expert opinion: Questions remain regarding the biological functions of FAP-α, either dependent or independent of its enzyme activity. However, as progress is underway to develop FAP-α-specific inhibitors and therapeutic antibodies, its role in diseases associated with fibrosis is starting to emerge, ultimately leading to novel therapeutic options for inflammatory and oncologic diseases.
Topics: Animals; Disease Progression; Drug Design; Endopeptidases; Fibroblasts; Fibrosis; Gelatinases; Gene Expression Regulation; Humans; Membrane Proteins; Molecular Targeted Therapy; Neoplasms; Serine Endopeptidases
PubMed: 28829211
DOI: 10.1080/14728222.2017.1370455 -
Current Biology : CB Apr 2002
Review
Topics: Animals; COP9 Signalosome Complex; Cysteine Endopeptidases; Multienzyme Complexes; Multiprotein Complexes; Peptide Hydrolases; Peptide Initiation Factors; Prokaryotic Initiation Factor-3; Proteasome Endopeptidase Complex; Proteins
PubMed: 11937035
DOI: 10.1016/s0960-9822(02)00775-3 -
American Journal of Hematology Dec 2002It is becoming clear that a number of proteins regulating cellular mechanisms for homeostasis in all eukaryotes are controlled not only by phosphorylation and... (Review)
Review
It is becoming clear that a number of proteins regulating cellular mechanisms for homeostasis in all eukaryotes are controlled not only by phosphorylation and dephosphorylation but also by ubiquitination and deubiquitination. This includes most of oncoproteins and signaling components involved in receptor tyrosine kinase (RTK)-mediated signal transduction pathways. Like protein phosphorylation and dephosphorylation regulated by kinases and phosphatases, respectively, protein ubiquitination and deubiquitination are very dynamic and are regulated by ubiquitin conjugating enzymes and deubiquitinating (DUB) enzymes. A number of deubiquitinating enzymes have been isolated even though little is known about their biological functions. This review concentrates on recent findings and new insights into DUB enzyme subfamily members in lymphocytes.
Topics: Amino Acid Sequence; Animals; Conserved Sequence; Cysteine Endopeptidases; Cytokines; Endopeptidases; Enzyme Induction; Humans; Immediate-Early Proteins; Lymphocytes; Mice; Molecular Sequence Data; Multienzyme Complexes; Proteasome Endopeptidase Complex; Sequence Alignment; Sequence Homology, Amino Acid
PubMed: 12447969
DOI: 10.1002/ajh.10243 -
Beijing Da Xue Xue Bao. Yi Xue Ban =... Oct 2009Osteoarthritis is mainly caused by the degenerative changes of cartilage and cartilage extracellular matrix, while Aggrecanases degradate Proteoglycans which are the... (Review)
Review
Osteoarthritis is mainly caused by the degenerative changes of cartilage and cartilage extracellular matrix, while Aggrecanases degradate Proteoglycans which are the major components of cartilage. This review includes three aspects: (1) We have concluded the major enzymes(ADAMTS-4 and ADAMTS-5) which regulate the metabolism of cartilage extracellular matrix. Meanwhile, we have summarized the structure of aggrecanases(ADAMTS-4 and ADAMTS-5) and introduced the function of each regional structure; (2) We have concluded the way cytokines and glycosaminoglycans regulate the metabolism of aggrecanases, and discussed the regulation and control principle of cytokines and glycosaminoglycan; (3) We have summarized the majority of inhibitors to the aggrecanases, introduced the endogenic inhibitors, and put our emphasis on the extrinsic inhibitors (chelating agents, polypeptides and so on). Through deeper research on the enzymes, it will help us further understand the pathogenesis of osteoarthritis, and open up new avenues to clinical treatment.
Topics: ADAM Proteins; ADAMTS4 Protein; ADAMTS5 Protein; Endopeptidases; Extracellular Matrix; Humans; Osteoarthritis; Procollagen N-Endopeptidase; Tissue Inhibitor of Metalloproteinase-3
PubMed: 19829688
DOI: No ID Found -
The FEBS Journal May 2017Proteases are enzymes that hydrolyze the peptide bond of peptide substrates and proteins. Despite significant progress in recent years, one of the greatest challenges in... (Review)
Review
Proteases are enzymes that hydrolyze the peptide bond of peptide substrates and proteins. Despite significant progress in recent years, one of the greatest challenges in the design and testing of substrates, inhibitors and activity-based probes for proteolytic enzymes is achieving specificity toward only one enzyme. This specificity is particularly important if the enzyme is present with other enzymes with a similar catalytic mechanism and substrate specificity but completely different functionality. The cross-reactivity of substrates, inhibitors and activity-based probes with other enzymes can significantly impair or even prevent investigations of a target protease. In this review, we describe important concepts and the latest challenges, focusing mainly on peptide-based substrate specificity techniques used to distinguish individual enzymes within major protease families.
Topics: Animals; Endopeptidases; Humans; Peptide Hydrolases; Protease Inhibitors; Substrate Specificity
PubMed: 28052575
DOI: 10.1111/febs.14001