Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: UNIPROT:P10415 (Bcl-2)
 33,771 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Bid is a key member of the Bcl-2 family proteins involved in the control of the apoptotic cascade in cells, leading to cell death. Uncontrolled cell death is associated with several human pathologies, such as neurodegenerative diseases and ischemic injuries. Therefore, Bid represents a potential yet unexplored and challenging target for strategies aimed at the development of therapeutic agents. Here we show that a multidisciplinary NMR-based approach that we named SAR by ILOEs (structure activity relationships by interligand nuclear Overhauser effect) allowed us to rationally design a series of 4-phenylsulfanyl-phenylamine derivatives that are capable of occupying a deep hydrophobic crevice on the surface of Bid. These compounds represent the first antiapoptotic small molecules targeting a Bcl-2 protein as shown by their ability to inhibit tBid-induced SMAC release, caspase-3 activation, and cell death.
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PMID:Targeting apoptosis via chemical design: inhibition of bid-induced cell death by small organic molecules. 1532 12

Rabdosia rubescens is a herbal medicine used to treat esophageal cancer in China. In this study, the sesquiterpene oridonin, an isoprenoid, was isolated from Rabdosia rubescens. Mass spectroscopy and carbon 13 NMR spectroscopy were used to identify the structure of the purified compound. It was then evaluated for biological activity against human cell lines derived from prostate (DU-145, LNCaP), breast (MCF-7), and ovarian (A2780 and PTX10) cancers. Oridonin exhibited anti-proliferative activity toward all cancer cell lines tested, with an IC50 estimated by the MTT cell viability assay ranging from 5.8+/-2.3 to 11.72+/-4.8 microM. Flow cytometric analysis demonstrated that oridonin induced a G1 phase arrest in androgen receptor-positive LNCaP cells containing wt p53, while it blocked the cell cycle at G2 and M phases in androgen receptor-negative DU-145 cells with mutated p53; the arrest in M was verified by examination of cell morphology and by the increased frequency of cells with Ser-10 phosphorylated histone H3. The increased incidence of apoptosis, identified by characteristic changes in cell morphology, was seen in tumor lines treated with oridonin. Notably, at concentrations that induced apoptosis among tumor cells, oridonin failed to induce apoptosis in cultures of normal human fibroblasts. Western blot analysis was used to determine the protein expression of cancer suppressor genes, p53 (wt) and Bax, and the proto-oncogene, Bcl-2 in LNCaP cells following treatment with oridonin. Oridonin up-regulated p53 and Bax and down-regulated Bcl-2 expression in a dose-dependent manner. To further explore the possible interaction between oridonin and DNA, its absorption spectrum was measured in the presence and absence of double stranded (ds) DNA. Spectral shifts and an increase in absorption band intensity were observed indicating interaction of oridonin with DNA bases. The nature of the binding is not clear at present though no evidence of histone H2AX phosphorylation on Ser-139 was apparent in DU-145 cells treated with oridonin that would indicate the induction of ds DNA breaks. In conclusion, oridonin inhibits cancer cell growth in a cell cycle specific manner and shifts the balance between pro- and anti-apoptotic proteins in favor of apoptosis. The present data suggest that further studies are warranted to assess the potential of oridonin in cancer prevention and/or treatment.
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PMID:The cytostatic and cytotoxic effects of oridonin (Rubescenin), a diterpenoid from Rabdosia rubescens, on tumor cells of different lineage. 1570 11

Bcl-2 contains an unusually long loop between the first and the second helices. This loop has been shown to be highly flexible based on NMR and X-ray crystallographic analyses of this region. Bcl-2 is regulated at the posttranslational level through phosphorylation of specific residues within the flexible loop. The biological role and posttranslational modifications of the loop of Bcl-2 is currently unclear. FK-506 binding protein 38 (FKBP38) has been reported to interact with Bcl-2, suggesting that FKBP38 could act as a docking molecule to localize Bcl-2 at the mitochondrial membrane [Shirane, M. and Nakayama, K.I. (2003) Inherent calcineurin inhibitor FKBP38 targets Bcl-2 to mitochondria and inhibits apoptosis. Nat. Cell Biol. 5, 28-37]. Here, we investigated the molecular interaction between FKBP38 and Bcl-2, and demonstrated that Bcl-2 interacts with FKBP38 through the unstructured loop, and the interaction appears to regulate phosphorylation in the loop of Bcl-2.
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PMID:The flexible loop of Bcl-2 is required for molecular interaction with immunosuppressant FK-506 binding protein 38 (FKBP38). 1573 59

Solid phase synthesis of BH4, the 26 amino-acid domain (6RTGYDNREIVMKYIHYKLSQRGYEWD31) of the anti-apoptotic Bcl-2 protein has been accomplished using Fmoc chemistry. The use of peculiar cleavage conditions provided high yields after purification such that tens to hundreds of mg could be obtained. A 15N-labelled version of the peptide could also be synthesized for NMR studies in membranes. The peptide purity was not lower than 98% as controlled by UV and MALDI-TOF mass spectrometry. The secondary structure was determined in water, trifluoroethanol (TFE) and in lipid membrane using UV circular dichroism. The peptide shows dominant beta-sheeted structures in water that convert progressively into alpha-helical features upon addition of TFE or membrane. The amphipathic character of the helix suggests that the peptide might have a structure akin to those of antimicrobial peptides upon interaction with membranes.
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PMID:Synthesis and secondary structure in membranes of the Bcl-2 anti-apoptotic domain BH4. 1594 41

Infrared spectroscopy was used to study the secondary structure of peptides which imitate the amino acid sequences of the C-terminal domains of the pro-apoptotic protein Bak (Bak-C) and the anti-apoptotic protein Bcl-2 (Bcl-2-C) when incorporated into different lipid vesicles. Whereas beta-pleated sheet was the predominant type of secondary structure of Bak-C in the absence of membranes, the same peptide adopted different structures depending on lipid composition when incorporated into membranes, with the predominance of the alpha-helical structure in the case of DMPC and other phospholipids, such as POPC and POPG. However, beta-pleated sheet was the predominant structure in other membranes containing phospholipids with longer fatty acyl chains and cholesterol, as well as in a mixture which imitates the composition of the outer mitochondrial membrane (OMM). Similarly, Bcl-2-C adopted a structure with a predominance of intermolecularly bound pleated beta-sheet in the absence of membranes, with alpha-helix as the main component in the presence of DMPC and POPG, but intermolecular beta-sheet in the presence of EYPC and cholesterol. Using ATR-IR, it was found that the orientation of the alpha-helical components of both domains was nearly perpendicular to the plane of the membrane in the presence of DMPC membranes, but not in EYPC or OMM membranes. (2)H NMR spectroscopy of DMPC-d(54) confirmed the transmembrane disposition of the domains, revealing that they broadened the phase transition temperature, although the order parameter of the C-D bonds was not affected, as might have been expected for intrinsic peptides. When all these results are taken together, it was concluded that the domains only form transmembrane helices in membranes of reduced thickness and that hydrophobic mismatching occurs in thicker membranes, as happens in the membrane imitating the composition of the OMM, where the peptides were partially located outside the membranes.
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PMID:Modulation of the membrane orientation and secondary structure of the C-terminal domains of Bak and Bcl-2 by lipids. 1608 82

The immunosuppressant FK-506 binding protein 38 (FKBP38) is localized at the mitochondrial membrane and appears to play an important role in apoptosis. Recent reports about the potential functions of FKBP38 in apoptosis appear to be controversial. To further understand the biological function of FKBP38, here, we studied its molecular characteristics and a potential regulatory role on the anti-apoptotic protein Bcl-2. Our results suggest that FKBP38 appears to show chaperone activities in the citrate synthase aggregation assays during thermal denaturation and affect solubility of Bcl-2 when they are co-expressed. The FKBP family proteins bind the immunosuppressive drug FK-506 through the FK-506 binding domain and consequently inhibit the activity of calcineurin. In this study, from our NMR studies and calcineurin assays in vitro, we demonstrate that the N-terminal fragment of FKBP38 which contains the FK-506 binding domain does not bind FK-506 at molecular level. Lastly, to investigate the effect of FKBP38 on Bcl-2, we suppressed FKBP38 by RNA interference (RNAi) of FKBP38. Our results suggest that the suppression of FKBP38 appears to make Bcl-2 unstable or unprotected from degradation in an unknown mechanism.
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PMID:Molecular characterization of FK-506 binding protein 38 and its potential regulatory role on the anti-apoptotic protein Bcl-2. 1617 96

The antiapoptotic proteins Bcl-x(L) and Bcl-2 play key roles in the maintenance of normal cellular homeostasis. However, their overexpression can lead to oncogenic transformation and is responsible for drug resistance in certain types of cancer. This makes Bcl-x(L) and Bcl-2 attractive targets for the development of potential anticancer agents. Here we describe the structure-based discovery of a potent Bcl-x(L) inhibitor directed at a hydrophobic groove on the surface of the protein. This groove represents the binding site for BH3 peptides from proapoptotic Bcl-2 family members such as Bak and Bad. Application of NMR-based screening yielded an initial biaryl acid with an affinity (K(d)) of approximately 300 microM for the protein. Following the classical "SAR by NMR" approach, a second-site ligand was identified that bound proximal to the first-site ligand in the hydrophobic groove. From NMR-based structural studies and parallel synthesis, a potent ligand was obtained, which binds to Bcl-x(L) with an inhibition constant (K(i)) of 36 +/- 2 nM.
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PMID:Discovery of a potent inhibitor of the antiapoptotic protein Bcl-xL from NMR and parallel synthesis. 1642 51

We report the first G-quadruplex structure formed in the promoter region of the human bcl-2. Bcl-2 is a potent oncoprotein that functions as an inhibitor of cell apoptosis and has been found to be aberrantly overexpressed in a wide range of human tumors. A highly GC-rich region upstream of the P1 promoter plays an important role in the regulation of the transcriptional activity of the bcl-2 oncogene. The purine-rich strand of this region contains multiple runs of guanines and can form three distinct intramolecular G-quadruplexes in K+-containing solution. Of these, the G-quadruplex formed within the middle four consecutive guanine runs has been shown to be the most stable G-quadruplex structure, while it is also a mixture of loop isomers. The predominant G-quadruplex structure formed in this region was studied by NMR. Our results demonstrate a novel folding of a unique intramolecular G-quadruplex structure with mixed parallel/antiparallel G-strands. This G-quadruplex structure contains three G-tetrads connected with a single-nucleotide double-chain-reversal side loop and two lateral loops. The first three-nucleotide CGC loop in the bcl-2 promoter sequence forms a lateral loop, as opposed to a double-chain-reversal side loop observed in a similar sequence in the c-MYC promoter, which appears to largely determine the overall folding of the bcl-2 G-quadruplex. Furthermore, both the bcl-2 and c-MYC promoter sequences contain the G3NG3 sequence motif, which forms a stable double-chain-reversal, parallel-stranded structural motif. This predominant bcl-2 G-quadruplex represents an attractive novel target for the design of new anticancer drugs that specifically modulate bcl-2 gene expression.
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PMID:An intramolecular G-quadruplex structure with mixed parallel/antiparallel G-strands formed in the human BCL-2 promoter region in solution. 1643 24

The functional genomic approaches of transcriptomics, proteomics and metabolomics aim to measure the mRNA, protein or metabolite complement of a cell, tissue or organism. In this study we have investigated the compatibility of transcriptional analysis, using Reverse Transcription (RT)-PCR, and metabolite analysis, by high-resolution magic angle spinning (HRMAS) 1H NMR spectroscopy, in BT4C rat glioma following the induction of programmed cell death. The metabolite and transcriptional changes that accompanied apoptosis were examined at 0, 4 and 8 days of ganciclovir/thymidine kinase gene therapy. Despite the high spinning speeds employed during HRMAS 1H NMR spectroscopy of one-half of the tumor samples, RT-PCR analysis of the pro-apoptotic transcripts Bcl-2, BAK-1, caspase-9 and FAS was possible, producing similar results to those detected in the unspun half of the tumors. Furthermore, the expression of FAS was inversely correlated with some of the key metabolic changes across the time period examined including the increases CH=CH and CH=CHCH2 lipid resonances which accompany apoptosis. This study demonstrates how combined transcriptomic and metabolomic studies of tumors can be used to understand the molecular events that accompany well documented metabolic perturbations during cell death processes.
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PMID:High-resolution magic angle spinning 1H NMR spectroscopy and reverse transcription-PCR analysis of apoptosis in a rat glioma. 1650 6

Isocostunolide is a sesquiterpene lactone isolated from the roots of Inula helenium. Its chemical structure was determined by NMR and FAB-MS spectra. No biological activities of this compound have yet been reported. In this study, we found isocostunolide could effectively induce cytotoxicity in three cancer cell lines (A2058, HT-29, and HepG2), with an IC(50) of 3.2, 5.0, and 2.0 micro g/mL, respectively. DNA flow cytometric analysis indicated that isocostunolide actively induced apoptosis of cancer cells accompanied by a marked loss of G0/G1 phase cells. To address the mechanism of the apoptotic effect of isocostunolide, we analyzed the induction of apoptosis-related proteins in A2058. The levels of pro-caspase-8, Bid, pro-caspase-3, and poly(ADP-ribose) polymerase (PARP) decreased. However, the level of Fas was increased markedly in a dose-dependent manner. Furthermore, this compound markedly induced a depolarization of mitochondrial membranes to facilitate cytochrome c release into cytosol. The findings suggest that isocostunolide may activate a mitochondria-mediated apoptosis pathway. To address this, we found that isocostunolide-induced loss of mitochondrial membrane potential occurred via modulation of the Bcl-2 family proteins. The production of intracellular reactive oxygen species (ROS) in A2058 was not elicited. In summary, for the first time, we have isolated and characterized isocostunolide from I. helenium. This compound induces apoptosis through a mitochondria-dependent pathway in A2058 cells.
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PMID:Isocostunolide, a sesquiterpene lactone, induces mitochondrial membrane depolarization and caspase-dependent apoptosis in human melanoma cells. 1669 6


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