Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.22.56 (caspase-3)
35,750 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We describe the characterization and purification of a protease that cleaves sterol regulatory element-binding protein-1 (SREBP-1) and SREBP-2 in vitro. Cleavage occurs between the basic helix-loop-helix-leucine zipper and the first transmembrane domain of each SREBP. This is the region in which the SREBPs are cleaved physiologically by a sterol-regulated protease that releases an NH2-terminal fragment that activates transcription of the genes for the low density lipoprotein receptor and 3-hydroxy-3-methylglutaryl CoA synthase. The cleavage enzyme, designated SREBP cleavage activity (SCA), belongs to a new class of cysteine proteases of the interleukin-1 beta-converting enzyme (ICE) family, all of which cleave at aspartic acid residues. Like ICE, SCA was inactive in cytosol, and it was activated in vitro by incubation at 30 degrees C. SCA was resistant to inhibitors of serine, aspartyl, and metalloproteases, but it was sensitive to N-ethylmaleimide. The enzyme cleaved SREBP-1 and SREBP-2 between the Asp and Ser of a conserved sequence (S/DEPDSP). The activity was blocked by a tetrapeptide aldehyde, Ac-Asp-Glu-Ala-Asp-aldehyde (Ac-DEAD-CHO). A purified preparation of SCA from hamster liver contained a prominent 20-kDa polypeptide that could be labeled with [14C]iodoacetic acid. Labeling was blocked by Ac-DEAD-CHO. Partial amino acid sequence of this polypeptide revealed that it was the hamster equivalent of human CPP32, a putative protease whose cDNA was recently identified by virtue of sequence homology to ICE. CPP32 and ICE have been implicated in apoptosis in animal cells. Whether SCA/CPP32 participates in vivo in the sterol-regulated activation of SREBP, or whether it activates SREBPs during apoptosis, remains to be determined.
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PMID:Purification of an interleukin-1 beta converting enzyme-related cysteine protease that cleaves sterol regulatory element-binding proteins between the leucine zipper and transmembrane domains. 762 13

We have developed a PCR approach to clone new apoptotic Ced-3/Ice-like cysteine protease genes. This approach uses degenerate oligonucleotides encoding the highly conserved pentapeptides QACRG and GSWFI that are present in all known apoptotic cysteine proteases. Using this approach, we have cloned a novel apoptotic gene from human Jurkat T lymphocytes. The new gene encodes a approximately 34-kilodalton protein that is highly homologous to human CPP32, Caenorhabditis elegans cell death protein CED-3, mammalian Ich-1 (Nedd2), and mammalian interleukin-1 beta converting enzyme. Because of its high homology to the C. elegans Ced-3 gene, we named the new gene mammalian Ced-3 homologue Mch2. Two Mch2 transcripts (Mch2 alpha, 1.7 kb; Mch2 beta, 1.4 kb) were detected in Jurkat T lymphocytes and other cell lines. We believe that the Mch2 alpha transcript encodes the full-length Mch2, whereas the Mch2 beta transcript encodes a shorter Mch2 isoform, probably as a result of alternative splicing. Like interleukin-1 beta converting enzyme and CPP32, recombinant Mch2 alpha, but not Mch2 beta, possesses protease activity, as determined by its ability to cleave the fluorogenic peptide DEVD-AMC. CPP32 and Mch2 alpha can also cleave poly(ADP-ribose) polymerase in vitro, suggesting that these enzymes participate in poly(ADP-ribose) polymerase cleavage observed during cellular apoptosis. In addition, overexpression of recombinant Mch2 alpha, but not Mch2 beta, induces apoptosis in Sf9 insect cells. Our data suggest that Mch2 is a Ced-3/interleukin-1 beta converting enzyme-like cysteine protease and could be another important mediator of apoptosis in mammalian cells.
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PMID:Mch2, a new member of the apoptotic Ced-3/Ice cysteine protease gene family. 779 96

Recent evidence suggests that mammalian cysteine proteases related to Caenorhabditis elegans CED-3 are key components of mammalian programmed cell death or apoptosis. We have shown recently that the CPP32 and Mch2 alpha cysteine proteases cleave the apoptotic markers poly(ADP-ribose) polymerase (PARP) and lamins, respectively. Here we report the cloning of a new Ced-3/interleukin 1 beta-converting enzyme-related gene, designated Mch3, that encodes a protein with the highest degree of homology to CPP32 compared to other family members. An alternatively spliced isoform, named Mch3 beta, was also identified. Bacterially expressed recombinant Mch3 has intrinsic autocatalytic/autoactivation activity. The specific activity of Mch3 alpha toward the peptide substrate DEVD-7-amino-4-methylcoumarin and PARP resembles that of CPP32. Like interleukin 1 beta-converting enzyme and CPP32, the active Mch3 alpha is made of two subunits derived from a precursor (proMch3 alpha). It was of interest that recombinant CPP32-p17 subunit can form an active heteromeric enzyme complex with recombinant Mch3 alpha-p12 subunit and vice versa, as determined by the ability of the heteromeric complexes to induce apoptosis in Sf9 cells. These data suggest that proMch3 alpha and proCPP32 can interact to form an active Mch3 alpha/CPP32 heteromeric complex. We also provide evidence that CPP32 can efficiently cleave proMch3 alpha, but not the opposite, suggesting that Mch3 alpha activation in vivo may depend in part on CPP32 activity. The high degree of conservation in structure and specific activity and the coexistence of Mch3 alpha and CPP32 in the same cell suggests that the PARP cleavage activity observed during apoptosis cannot solely be attributed to CPP32 but could also be an activity of Mch3 alpha.
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PMID:Mch3, a novel human apoptotic cysteine protease highly related to CPP32. 852 91

Cysteine proteases of the interleukin 1 beta Converting Enzyme (ICE)/CED-3 family have been implicated in the effector process of apoptosis in several systems, including Fas-mediated apoptosis. We have recently isolated and partially characterized a protease present in extracts from anti-Fas antibody treated Jurkat T cells that promotes apoptotic changes in isolated nuclei (Schlegel, J., Peters, I., and Orrenius, S. (1995) FEBS Lett. 364, 139-142). We now show that this protease cleaves poly-(ADP-ribose) polymerase (PARP) with high efficiency and specificity. Both PARP proteolysis and the proapoptotic effects of the protease are inhibited by nanomolar concentrations of a selective inhibitor of apopain (CPP32), while an inhibitor of IL-1 beta converting enzyme is much less effective, requiring micromolar concentrations for the inhibition of the isolated protease. Kinetic analysis of the isolated protease reveals kinetic constants similar to those reported for apopain. The isolated protease is recognized by antibodies specific for CPP32/apopain but not by an anti-ICE antibody. Furthermore, a selective inhibitor of apopain prevents Fas-induced apoptosis in intact Jurkat T cells. We therefore conclude that CPP32/apopain is activated in Fas-induced apoptosis.
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PMID:CPP32/apopain is a key interleukin 1 beta converting enzyme-like protease involved in Fas-mediated apoptosis. 856 26

Genetic analyses of Caenorhabditis elegans has identified three genes that function in the regulation of nematode cell death. Mammalian homologs of two of these genes, ced-9 and ced-3, have been identified and comprise proteins belonging to the Bcl-2 and ICE families, respectively. To date, it is unclear where the negative regulators, ced-9 and bcl-2, function relative to the death effectors, ced-3 and the mammalian ced-3 homologs, respectively. Here, the molecular order of the cell death pathway is defined. Our results establish that Bcl-2 and Bcl-xL function upstream of two members of the ICE/CED-3 family of cysteine proteases, Yama (CPP32/apopain) and ICE-LAP3 (Mch3).
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PMID:Molecular ordering of the cell death pathway. Bcl-2 and Bcl-xL function upstream of the CED-3-like apoptotic proteases. 861 12

Apoptosis (programmed cell death) is a fundamental process for normal development of multicellular organisms, and is involved in the regulation of the immune system, normal morphogenesis, and maintenance of homeostasis, ICE/CED-3 family cysteine proteases have been implicated directly in apoptosis, but relatively few of the substrates through which their action is mediated have been identified. Here we report that D4-GDI, an abundant hematopoietic cell GDP dissociation inhibitor for the Ras-related Rho family GTPases, is a substrate of the apoptosis protease CPP32/Yama/Apopain. D4-GDI was rapidly truncated to a 23-kDa fragment in Jurkat cells with kinetics that parallel the onset of apoptosis following Fas cross-linking with agonistic antibody or treatment with staurosporine. Fas- and staurosporine-induced apoptosis as well as cleavage of D4-GDI were inhibited by the ICE inhibitor, YVAD-cmk. D4-GDI was cleaved in vitro by recombinant CPP32 expressed in Escherichia coli to form a 23-kDa fragment. The CPP32-mediated cleavage of D4-GDI was completely inhibited by 1 microM DEVD-CHO, a reported selective inhibitor of CPP32. In contrast, the ICE-selective inhibitors, YVAD-CHO or YVAD-cmk, did not inhibit CPP32-mediated D4-GDI cleavage at concentrations up to 50 microM. N-terminal sequencing of the 23-kDa D4-GDI fragment demonstrated that D4-GDI was cleaved between Asp19 and Ser20 of the poly(ADP-ribose) polymerase-like cleavage sequence DELD19S. These data suggest that regulation by D4-GDI of Rho family GTPases may be disrupted during apoptosis by CPP32-mediated cleavage of the GDI protein.
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PMID:D4-GDI, a substrate of CPP32, is proteolyzed during Fas-induced apoptosis. 862 69

Granzyme B plays an essential role in cytotoxic T lymphocyte (CTL)-mediated cell killing. Recent studies suggest that granzyme B may exert its effect by cleaving and activating CPP32, a member of the interleukin-1 beta-converting enzyme/Ced-3 family of cysteine proteases. We have examined the processing and activation of CMH-1, a close homologue of CPP32, by granzyme B in vitro. We have found that granzyme B specifically cleaves CMH-1 at Asp198-Ser199 between the p20 and p12 and activates the cysteine protease. Cleavage between p20 and the prosequence of CMH-1 at Asp23-Ala24 is autocatalytic and is not required for CMH-1 activity in vitro. The cleavage and activation of CMH-1 by granzyme B in vitro sugge st that, in addition to CPP32, CMH-1 may also play a role in CTL-mediated cell killing.
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PMID:Processing and activation of CMH-1 by granzyme B. 863 95

The new and growing family of interleukin-1beta-converting enzyme (ICE) cysteine proteases are now recognised to be major effectors of cellular death by apoptosis. Like other members of this family, the CPP32/Yama proform is activated by processing to its active heterodimeric enzyme or apopain when it likely contributes to the process of apoptosis by cleaving poly(ADP-ribose) polymerase (PARP) and thereby inhibiting much of its DNA repair activity. Apoptosis plays a fundamental role in the regulation of the immune system where it is involved in the selection of both T and B lymphocytes bearing antigen receptor (AgR) for non-self. Cells of the Ramos Epstein-Barr virus (EBV)-genome-negative Burkitt lymphoma (BL) B cell line (Ramos-BL) can be triggered into growth arrest and apoptosis by treating with the calcium ionophore ionomycin or by crosslinking their surface AgR with antibodies directed against immunoglobulin (Ig)M (anti-IgM). Ionomycin- and AgR-triggered growth arrest and apoptosis are arrested by signals transduced through the surface CD40 of Ramos-BL B cells. Both ionomycin and anti-IgM trigger activation of CPP32 and cleavage of PARP prior to the onset of apoptosis; this process is abrogated by treatment with anti-CD40 and is independent of Bcl-2 expression. A tripeptide inhibitor of ICE family cysteine proteases, Z-Val-Ala-Asp-fluoromethylketone (zVAD-fmk) inhibits ionomycin- and AgR-triggered CPP32 activation, PARP cleavage and apoptosis, but not growth arrest, in Ramos-BL B cells. Thus, in this report we demonstrate that in a physiological system, activation of endogenous members of the ICE family, including CPP32, and cleavage of the death substrate PARP act as major effectors of apoptotic death.
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PMID:Ligation of CD40 rescues Ramos-Burkitt lymphoma B cells from calcium ionophore- and antigen receptor-triggered apoptosis by inhibiting activation of the cysteine protease CPP32/Yama and cleavage of its substrate PARP. 864 64

The Bcl-2 family and the ICE family of cysteine proteases play important roles in regulating cell death. We show here that induction of cell death by a Ca2+ ionophore or hypoxia results in increased levels and activity of active ICE(-like) proteases and the subsequent activation of CPP32/Yama(-like) proteases, and that inhibition of these protease activities reduces the extent of cell death. Overexpression of the anti-apoptotic proteins Bcl-2 or Bcl-xL inhibits the cell death and the activation of ICE(-like) and CPP32/Yama(-like) proteases, indicating that Bcl-2 and Bcl-xL act upstream of these proteases. We also show that specific inhibition of ICE(-like) proteases in vivo prevents activation of CPP32/Yama(-like) proteases, whereas inhibition of CPP32/Yama(-like) proteases does not prevent activation of ICE(-like) proteases, suggesting the existence of a protease cascade in vivo that requires ICE(-like) proteases for activation of CPP32/Yama(-like) proteases. Induction of necrotic cell death by KCN also induces activation of ICE(-like) proteases but not of CPP32/Yama(-like) proteases, and Bcl-2 and Bcl-xL inhibit the activation and the cell death, suggesting that the functional site of Bcl-2 and Bcl-xL is also upstream of ICE(-like) proteases in at least some forms of necrosis.
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PMID:Bcl-2 expression prevents activation of the ICE protease cascade. 864 64

The Caenorhabditis elegans cell-death gene ced-3 encodes a protein similar to mammalian interleukin-1beta-converting enzyme (ICE), a cysteine protease implicated in mammalian apoptosis. We show that the full-length CED-3 protein undergoes proteolytic activation to generate a CED-3 cysteine protease and that CED-3 protease activity is required for killing cells by programmed cell death in C. elegans. We developed an easy and general method for the purification of CED-3/ICE-like proteases and used this method to facilitate a comparison of the substrate specificities of four different purified cysteine proteases. We found that in its substrate preferences CED-3 was more similar to the mammalian CPP32 protease than to mammalian ICE or NEDD2/ICH-1 protease. Our results suggest that different mammalian CED-3/ICE-like proteases may have distinct roles in mammalian apoptosis and that CPP32 is a candidate for being a mammalian functional equivalent of CED-3.
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PMID:The Caenorhabditis elegans cell-death protein CED-3 is a cysteine protease with substrate specificities similar to those of the human CPP32 protease. 865 23


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