Secondary literature sources for DWNN
The following references were automatically generated.
- Moela P, Choene MM, Motadi LR
- Silencing RBBP6 (Retinoblastoma Binding Protein 6) sensitises breast cancer cells MCF7 to staurosporine and camptothecin-induced cell death.
- Immunobiology. 2014; 219: 593-601
- Display abstract
Retinoblastoma Binding Protein 6 (RBBP6) is a multi-domain protein that uses its ring finger domain to interact with p53 and pRb tumour suppressor genes. The mechanism by which RBBP6 uses to degrade p53 is still unknown; nonetheless it is well known that RBBP6 promotes cell proliferation in several cancers by negatively regulating p53 via its E3 ubiquitin ligase activity. Degradation of p53 by RBBP6 may compromise p53-mediated apoptosis in breast cancer. This study is intended to investigate, the potential applications of RNA interference (RNAi) to block RBBP6 expression, as well as its subsequent effect on cell growth and apoptosis. Our studies indicate that the knockdown of RBBP6 by siRNA modulates p53 gene expression involved in cell death pathways and apoptosis, showing statistically significant gene expression differences. RBBP6 siRNA significantly reduced cell growth compared to the control samples and inhibition of cellular proliferation was observed between 24 and 48h, as shown in the data obtained by real time cell analysis using the xCELLigence system. These results were further confirmed by flow cytometer which showed some apoptotic activity. About 20.7% increase in apoptosis was observed in cells co-treated with RBBP6 siRNA and camptothecin when compared to camptothecin-only whereas in siRBBP6 and staurosporine treated cells there was only an 8.8% increase in apoptosis. These findings suggest that silencing RBBP6 may be a novel strategy to promote camptothecin-induced apoptosis in breast cancer cells.
- Rizzo AA, Salerno PE, Bezsonova I, Korzhnev DM
- NMR structure of the human Rad18 zinc finger in complex with ubiquitin defines a class of UBZ domains in proteins linked to the DNA damage response.
- Biochemistry. 2014; 53: 5895-906
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Ubiquitin-mediated interactions are critical for the cellular DNA damage response (DDR). Therefore, many DDR-related proteins contain ubiquitin-binding domains, including ubiquitin-binding zinc fingers (UBZs). The majority of these UBZ domains belong to the C2H2 (type 3 Poleta-like) or C2HC (type 4 Rad18-like) family. We have used nuclear magnetic resonance (NMR) spectroscopy to characterize the binding to ubiquitin and determine the structure of the type 4 UBZ domain (UBZ4) from human Rad18, which is a key ubiquitin ligase in the DNA damage tolerance pathway responsible for monoubiquitination of the DNA sliding clamp PCNA. The Rad18-UBZ domain binds ubiquitin with micromolar affinity and adopts a beta1-beta2-alpha fold similar to the previously characterized type 3 UBZ domain (UBZ3) from the translesion synthesis DNA polymerase Poleta. However, despite nearly identical structures, a disparity in the location of binding-induced NMR chemical shift perturbations shows that the Rad18-UBZ4 and Poleta-UBZ3 domains bind ubiquitin in distinctly different modes. The Rad18-UBZ4 domain interacts with ubiquitin with the alpha-helix and strand beta1 as shown by the structure of the Rad18-UBZ domain-ubiquitin complex determined in this work, while the Poleta-UBZ3 domain exclusively utilizes the alpha-helix. Our findings suggest the existence of two classes of UBZ domains in DDR-related proteins with similar structures but unique ubiquitin binding properties and provide context for further study to establish the differential roles of these domains in the complex cellular response to DNA damage.
- Daniel J, Liebau E
- The ufm1 cascade.
- Cells. 2014; 3: 627-38
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The ubiquitin-fold modifier 1 (Ufm1) is a posttranslational modifier that belongs to the ubiquitin-like protein (UBL) family. Ufm1 is present in nearly all eukaryotic organisms, with the exception of fungi. It resembles ubiquitin in its ability to be ligated to other proteins, as well as in the mechanism of ligation. While the Ufm1 cascade has been implicated in endoplasmic reticulum functions and cell cycle control, its biological role still remains poorly understood. In this short review, we summarize the current state of Ufm1 research and its potential role in human diseases, like diabetes, ischemic heart disease and cancer.
- Rimsa V, Eadsforth TC, Hunter WN
- Structure of the SCAN domain of human paternally expressed gene 3 protein.
- PLoS One. 2013; 8: 69538-69538
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Human paternally expressed gene 3 protein (PEG3) is a large multi-domain entity with diverse biological functions, including acting as a transcription factor. PEG3 contains twelve Cys2-His2 type zinc finger domains, extended regions of predicted disorder and at the N-terminus a SCAN domain. PEG3 has been identified as partner of the E3 ubiquitin-protein ligase Siah1, an association we sought to investigate. An efficient bacterial recombinant expression system of the human PEG3-SCAN domain was prepared and crystals appeared spontaneously when the protein was being concentrated after purification. The structure was determined at 1.95 A resolution and reveals a polypeptide fold of five helices in an extended configuration. An extensive dimerization interface, using almost a quarter of the solvent accessible surface, and key salt bridge interactions explain the stability of the dimer. Comparison with other SCAN domains reveals a high degree of conservation involving residues that contribute to the dimer interface. The PEG3-SCAN domain appears to constitute an assembly block, enabling PEG3 homo- or heterodimerization to control gene expression in a combinatorial fashion.
- Motadi LR, Bhoola KD, Dlamini Z
- Expression and function of retinoblastoma binding protein 6 (RBBP6) in human lung cancer.
- Immunobiology. 2011; 216: 1065-73
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Retinoblastoma binding protein 6 (RBBP6) interacts with both p53 and pRb, and has been identified as an E3 ubiquitin ligase due to the presence of a RING finger domain. RBBP6 promotes the degradation of p53, thereby increasing cell proliferation. However it is not known whether RBBP6 is expressed in lung cancer, or interacts with p53 and pRB to modulate the proliferation or apoptosis of lung cancer cells. As assessed by immunohistochemistry, RBBP6 and p53 proteins were overexpressed in lung adenocarcinomas and lung squamous cell carcinomas. Expression of RBBP6 mRNA in lung tumor tissue was demonstrated by quantitative RT-PCR and fluorescence in situ hybridization. Expression of RBBP6 mRNA was low in poorly differentiated tumors but high in well-differentiated tumors. Bronchoalveolar carcinomas showed intense RBBP6 mRNA hybridization in the cytoplasm of cells undergoing mitosis, supporting the association between RBBP6 expression and cell proliferation. By qRT-PCR, RBBP6 mRNA expression was 1.6 fold higher, whereas p53 mRNA expression was 2.9 fold lower in lung tumors compared with normal lung tissue. Transfection of lung adenocarcinoma and squamous cell carcinoma cells with RBBP6 siRNA decreased RBBP6 mRNA expression, whereas transfection with p53 siRNA increased RBBP6 mRNA expression. Treatment with RBBP6 siRNA increased the ratio of Bax/Bcl2 mRNA, suggesting that RBBP6 may have an anti-apoptotic function in lung cancer cells. This is the first report that RBBP6 mRNA and "its protein products" are expressed in subtypes of human lung cancer. RBBP6 may be involved in the degradation of p53, thereby enhancing cell proliferation and inhibiting apoptosis in lung cancer.
- Bessiere D et al.
- Structure-function analysis of the THAP zinc finger of THAP1, a large C2CH DNA-binding module linked to Rb/E2F pathways.
- J Biol Chem. 2008; 283: 4352-63
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THAP1, the founding member of a previously uncharacterized large family of cellular proteins (THAP proteins), is a sequence-specific DNA-binding factor that has recently been shown to regulate cell proliferation through modulation of pRb/E2F cell cycle target genes. THAP1 shares its DNA-binding THAP zinc finger domain with Drosophila P element transposase, zebrafish E2F6, and several nematode proteins interacting genetically with the retinoblastoma protein pRb. In this study, we report the three-dimensional structure and structure-function relationships of the THAP zinc finger of human THAP1. Deletion mutagenesis and multidimensional NMR spectroscopy revealed that the THAP domain of THAP1 is an atypical zinc finger of approximately 80 residues, distinguished by the presence between the C2CH zinc coordinating residues of a short antiparallel beta-sheet interspersed by a long loop-helix-loop insertion. Alanine scanning mutagenesis of this loop-helix-loop motif resulted in the identification of a number of critical residues for DNA recognition. NMR chemical shift perturbation analysis was used to further characterize the residues involved in DNA binding. The combination of the mutagenesis and NMR data allowed the mapping of the DNA binding interface of the THAP zinc finger to a highly positively charged area harboring multiple lysine and arginine residues. Together, these data represent the first structure-function analysis of a functional THAP domain, with demonstrated sequence-specific DNA binding activity. They also provide a structural framework for understanding DNA recognition by this atypical zinc finger, which defines a novel family of cellular factors linked to cell proliferation and pRb/E2F cell cycle pathways in humans, fish, and nematodes.
- Burroughs AM, Balaji S, Iyer LM, Aravind L
- Small but versatile: the extraordinary functional and structural diversity of the beta-grasp fold.
- Biol Direct. 2007; 2: 18-18
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BACKGROUND: The beta-grasp fold (beta-GF), prototyped by ubiquitin (UB), has been recruited for a strikingly diverse range of biochemical functions. These functions include providing a scaffold for different enzymatic active sites (e.g. NUDIX phosphohydrolases) and iron-sulfur clusters, RNA-soluble-ligand and co-factor-binding, sulfur transfer, adaptor functions in signaling, assembly of macromolecular complexes and post-translational protein modification. To understand the basis for the functional versatility of this small fold we undertook a comprehensive sequence-structure analysis of the fold and developed a natural classification for its members. RESULTS: As a result we were able to define the core distinguishing features of the fold and numerous elaborations, including several previously unrecognized variants. Systematic analysis of all known interactions of the fold showed that its manifold functional abilities arise primarily from the prominent beta-sheet, which provides an exposed surface for diverse interactions or additionally, by forming open barrel-like structures. We show that in the beta-GF both enzymatic activities and the binding of diverse co-factors (e.g. molybdopterin) have independently evolved on at least three occasions each, and iron-sulfur-cluster-binding on at least two independent occasions. Our analysis identified multiple previously unknown large monophyletic assemblages within the beta-GF, including one which unifies versions found in the fasciclin-1 superfamily, the ribosomal protein L25, the phosphoribosyl AMP cyclohydrolase (HisI) and glutamine synthetase. We also uncovered several new groups of beta-GF domains including a domain found in bacterial flagellar and fimbrial assembly components, and 5 new UB-like domains in the eukaryotes. CONCLUSION: Evolutionary reconstruction indicates that the beta-GF had differentiated into at least 7 distinct lineages by the time of the last universal common ancestor of all extant organisms, encompassing much of the structural diversity observed in extant versions of the fold. The earliest beta-GF members were probably involved in RNA metabolism and subsequently radiated into various functional niches. Most of the structural diversification occurred in the prokaryotes, whereas the eukaryotic phase was mainly marked by a specific expansion of the ubiquitin-like beta-GF members. The eukaryotic UB superfamily diversified into at least 67 distinct families, of which at least 19-20 families were already present in the eukaryotic common ancestor, including several protein and one lipid conjugated forms. Another key aspect of the eukaryotic phase of evolution of the beta-GF was the dramatic increase in domain architectural complexity of proteins related to the expansion of UB-like domains in numerous adaptor roles.
- Chen D, Gould C, Garza R, Gao T, Hampton RY, Newton AC
- Amplitude control of protein kinase C by RINCK, a novel E3 ubiquitin ligase.
- J Biol Chem. 2007; 282: 33776-87
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Protein kinase C (PKC) isozymes play a central role in cellular signaling. Levels of PKC control the amplitude of agonist-induced signaling and alterations in these levels are associated with disease states, most notably cancer, yet mechanisms that control the turnover of the protein are poorly understood. Here we identify an E3 ligase that catalyzes the ubiquitin-mediated degradation of PKC. Specifically, we identified a RING finger domain-containing protein, RINCK (for RING-finger protein that interacts with C kinase) from a yeast two-hybrid screen using the amino terminus of PKCbeta as bait. RINCK encodes a protein of 581 amino acids that contains a RING finger domain, a B-box, and two coiled-coil regions, the three domains that form the signature motif of the large family of diverse TRIM (tripartite motif) proteins. Co-immunoprecipitation studies using tsA201 cells reveal that RINCK and PKC associate with each other in cells. Studies using fragments of PKCbeta reveal that this interaction is mediated by the C1A domain of PKC. RINCK induces the ubiquitination of PKC both in vitro and in cells. Overexpression of RINCK reduces the levels of PKC in cells, whereas genetic knockdown of endogenous RINCK increases the levels of PKC. This increase was observed for all PKC isozymes examined (including conventional, novel, and atypical). The RINCK-mediated degradation of PKC occurs independently of the classic phorbol ester-mediated down-regulation: genetic depletion of RINCK had no effect on the phorbol ester-mediated down-regulation and, additionally, up-regulated the levels of isozymes that cannot bind phorbol esters. Our data reveal a novel mechanism that provides amplitude control in PKC signaling through ubiquitination catalyzed by RINCK, an E3 ligase that specifically recognizes the C1 domain of PKC isoforms.
- Allen MD, Bycroft M
- The solution structure of the ZnF UBP domain of USP33/VDU1.
- Protein Sci. 2007; 16: 2072-5
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USP33/VDU1 is a deubiquitinating enzyme that binds to the von Hippel-Lindau tumor suppressor protein. It also regulates thyroid hormone activation by deubiquitinating type 2 iodothyronine deiodinase. USP33/VDU1 contains a ZF UBP domain, a protein module found in many proteins in the ubiquitin-proteasome system. Several ZF UBP domains have been shown to bind ubiquitin, and a structure of a complex of the ZF UBP domain of isoT/USP5 and ubiquitin is available. In the present work, the solution structure of the ZF UBP domain of USP33/VDU1 has been determined by NMR spectroscopy. The structure differs from that of the USP5 domain, which contains only one of the three Zn ions present in the USP33/VDU1 structure. The USP33/VDU1 ZnF UBP domain does not bind to ubiquitin.
- Pai MT et al.
- Solution structure of the Ubp-M BUZ domain, a highly specific protein module that recognizes the C-terminal tail of free ubiquitin.
- J Mol Biol. 2007; 370: 290-302
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The BUZ/Znf-UBP domain is a distinct ubiquitin-binding module found in the cytoplasmic deacetylase HDAC6, the E3 ubiquitin ligase BRAP2/IMP, and a subfamily of deubiquitinating enzymes. Here, we report the solution structure of the BUZ domain of Ubp-M, a ubiquitin-specific protease, and its interaction with ubiquitin. Unlike the BUZ domain from isopeptidase T (isoT) that contains a single zinc finger, the Ubp-M BUZ domain features three zinc-binding sites consisting of 12 residues. These zinc ligands form a pair of cross-braced ring fingers encapsulated within a third zinc finger in the primary structure. In contrast to isoT, which can form an N-terminal loop swapped dimer in the crystal state, the formation of additional zinc fingers in the Ubp-M BUZ domain restricts its N-terminal loop to intra-domain interactions. The ubiquitin-binding site of the Ubp-M BUZ domain is mapped to the highly conserved, concave surface formed by the alpha 3 helix and the central beta-sheet. We further show that this site binds to the C-terminal tail of free ubiquitin, and corresponding peptides display essentially the same binding affinities as full-length ubiquitin does for the Ubp-M BUZ domain. However, modification of the G76(Ub) carboxylate group either by a peptide or isopeptide bond abolishes BUZ-domain interaction. The unique ubiquitin-recognition mode of the BUZ domain family suggests that they may function as "sensors" of free ubiquitin in cells to achieve regulatory roles in many aspects of ubiquitin-dependent processes.
- Angers S, Li T, Yi X, MacCoss MJ, Moon RT, Zheng N
- Molecular architecture and assembly of the DDB1-CUL4A ubiquitin ligase machinery.
- Nature. 2006; 443: 590-3
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Protein ubiquitination is a common form of post-translational modification that regulates a broad spectrum of protein substrates in diverse cellular pathways. Through a three-enzyme (E1-E2-E3) cascade, the attachment of ubiquitin to proteins is catalysed by the E3 ubiquitin ligase, which is best represented by the superfamily of the cullin-RING complexes. Conserved from yeast to human, the DDB1-CUL4-ROC1 complex is a recently identified cullin-RING ubiquitin ligase, which regulates DNA repair, DNA replication and transcription, and can also be subverted by pathogenic viruses to benefit viral infection. Lacking a canonical SKP1-like cullin adaptor and a defined substrate recruitment module, how the DDB1-CUL4-ROC1 E3 apparatus is assembled for ubiquitinating various substrates remains unclear. Here we present crystallographic analyses of the virally hijacked form of the human DDB1-CUL4A-ROC1 machinery, which show that DDB1 uses one beta-propeller domain for cullin scaffold binding and a variably attached separate double-beta-propeller fold for substrate presentation. Through tandem-affinity purification of human DDB1 and CUL4A complexes followed by mass spectrometry analysis, we then identify a novel family of WD40-repeat proteins, which directly bind to the double-propeller fold of DDB1 and serve as the substrate-recruiting module of the E3. Together, our structural and proteomic results reveal the structural mechanisms and molecular logic underlying the assembly and versatility of a new family of cullin-RING E3 complexes.
- Tettamanzi MC, Yu C, Bogan JS, Hodsdon ME
- Solution structure and backbone dynamics of an N-terminal ubiquitin-like domain in the GLUT4-regulating protein, TUG.
- Protein Sci. 2006; 15: 498-508
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The GLUT4-regulating protein, TUG, functions to retain GLUT4-containing membrane vesicles intracellularly and, in response to insulin stimulation, releases these vesicles to the cellular exocytic machinery for translocation to the plasma membrane. As part of our on going effort to describe the molecular basis for TUG function, we have determined the tertiary structure and characterized the backbone dynamics for an N-terminal ubiquitin-like domain (TUG-UBL1) using NMR spectroscopy. A well-ordered conformation is observed for residues 10-83 of full-length TUG, and confirms a beta-grasp or ubiquitin-like topology. Although not required for in vitro association with GLUT4, the functional role of the TUG-UBL1 domain has not yet been described. We undertook a limited literature review of similar N-terminal UBL domains and noted that a majority participate in protein-protein interactions, generally functioning as adaptor modules to physically associate the over all activity of the protein with a specific cellular process, such as the ubiquitin-proteasome pathway. In consistent fashion, TUG-UBL1 is not expected to participate in a covalent protein modification reaction as it lacks the characteristic C-terminal diglycine ("GG") motif required for conjugation to an acceptor lysine, and also lacks the three most common acceptor lysine residues involved in polyubiquitination. Additionally, analysis of the TUG-UBL1 molecular surface reveals a lack of conservation of the "Ile-44 hydrophobic face" typically involved in ubiquitin recognition. Instead, we speculate on the possible significance of a concentrated area of negative electrostatic potential with increased backbone mobility, both of which are features suggestive of a potential protein-protein interaction site.
- Nederveen AJ et al.
- RECOORD: a recalculated coordinate database of 500+ proteins from the PDB using restraints from the BioMagResBank.
- Proteins. 2005; 59: 662-72
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State-of-the-art methods based on CNS and CYANA were used to recalculate the nuclear magnetic resonance (NMR) solution structures of 500+ proteins for which coordinates and NMR restraints are available from the Protein Data Bank. Curated restraints were obtained from the BioMagResBank FRED database. Although the original NMR structures were determined by various methods, they all were recalculated by CNS and CYANA and refined subsequently by restrained molecular dynamics (CNS) in a hydrated environment. We present an extensive analysis of the results, in terms of various quality indicators generated by PROCHECK and WHAT_CHECK. On average, the quality indicators for packing and Ramachandran appearance moved one standard deviation closer to the mean of the reference database. The structural quality of the recalculated structures is discussed in relation to various parameters, including number of restraints per residue, NOE completeness and positional root mean square deviation (RMSD). Correlations between pairs of these quality indicators were generally low; for example, there is a weak correlation between the number of restraints per residue and the Ramachandran appearance according to WHAT_CHECK (r = 0.31). The set of recalculated coordinates constitutes a unified database of protein structures in which potential user- and software-dependent biases have been kept as small as possible. The database can be used by the structural biology community for further development of calculation protocols, validation tools, structure-based statistical approaches and modeling. The RECOORD database of recalculated structures is publicly available from http://www.ebi.ac.uk/msd/recoord.
- Mather A, Rakgotho M, Ntwasa M
- SNAMA, a novel protein with a DWNN domain and a RING finger-like motif: a possible role in apoptosis.
- Biochim Biophys Acta. 2005; 1727: 169-76
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We have characterized SNAMA a hitherto uncharacterized Drosophila protein that appears to play a role in apoptosis. SNAMA (something that sticks like glue) is a 1231 amino acid protein with a conserved 76 residue N-terminal domain called Domain With No Name (DWNN). The DWNN domain was first identified in cytotoxic T Cell-resistant CHO cells using promoter trap mutagenesis to screen for genes involved in apoptosis. Subsequently, this domain was identified in other eukaryotic organisms including animals and plants. The SNAMA transcript is abundant early in embryogenesis but reduced in older embryos and in adult males and females. Human and mouse homologues of SNAMA are known to bind to p53 and to the retinoblastoma protein (Rb) suggesting a role in transcriptional regulation and cell cycle control. We took advantage of a P-element insertion line in which the P-element is inserted in the first intron, to investigate the biological function of the gene. These mutants are lethal when homozygous. Apoptosis appears early during embryogenesis and is observed virtually throughout the gastrula. The DWNN domain has a ubiquitin-like fold and may interact with a subset of cellular proteins. There is also a conserved RING finger-like motif along the sequence of SNAMA following a C2HC zinc finger.
- Singh S, Tonelli M, Tyler RC, Bahrami A, Lee MS, Markley JL
- Three-dimensional structure of the AAH26994.1 protein from Mus musculus, a putative eukaryotic Urm1.
- Protein Sci. 2005; 14: 2095-102
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We have used NMR spectroscopy to determine the solution structure of protein AAH26994.1 from Mus musculus and propose that it represents the first three-dimensional structure of a ubiquitin-related modifier 1 (Urm1) protein. Amino acid sequence comparisons indicate that AAH26994.1 belongs to the Urm1 family of ubiquitin-like modifier proteins. The best characterized member of this family has been shown to be involved in nutrient sensing, invasive growth, and budding in yeast. Proteins in this family have only a weak sequence similarity to ubiquitin, and the structure of AAH26994.1 showed a much closer resemblance to MoaD subunits of molybdopterin synthases (known structures are of three bacterial MoaD proteins with 14%-26% sequence identity to AAH26994.1). The structures of AAH26994.1 and the MoaD proteins each contain the signature ubiquitin secondary structure fold, but all differ from ubiquitin largely in regions outside of this fold. This structural similarity bolsters the hypothesis that ubiquitin and ubiquitin-related proteins evolved from a protein-based sulfide donor system of the molybdopterin synthase type.
- Furukawa M, Xiong Y
- BTB protein Keap1 targets antioxidant transcription factor Nrf2 for ubiquitination by the Cullin 3-Roc1 ligase.
- Mol Cell Biol. 2005; 25: 162-71
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The concentrations and functions of many eukaryotic proteins are regulated by the ubiquitin pathway, which consists of ubiquitin activation (E1), conjugation (E2), and ligation (E3). Cullins are a family of evolutionarily conserved proteins that assemble by far the largest family of E3 ligase complexes. Cullins, via a conserved C-terminal domain, bind with the RING finger protein Roc1 to recruit the catalytic function of E2. Via a distinct N-terminal domain, individual cullins bind to a protein motif present in multiple proteins to recruit specific substrates. Cullin 3 (Cul3), but not other cullins, binds directly with BTB domains to constitute a potentially large number of BTB-CUL3-ROC1 E3 ubiquitin ligases. Here we report that the human BTB-Kelch protein Keap1, a negative regulator of the antioxidative transcription factor Nrf2, binds to CUL3 and Nrf2 via its BTB and Kelch domains, respectively. The KEAP1-CUL3-ROC1 complex promoted NRF2 ubiquitination in vitro and knocking down Keap1 or CUL3 by short interfering RNA resulted in NRF2 protein accumulation in vivo. We suggest that Keap1 negatively regulates Nrf2 function in part by targeting Nrf2 for ubiquitination by the CUL3-ROC1 ligase and subsequent degradation by the proteasome. Blocking NRF2 degradation in cells expressing both KEAP1 and NRF2 by either inhibiting the proteasome activity or knocking down Cul3, resulted in NRF2 accumulation in the cytoplasm. These results may reconcile previously observed cytoplasmic sequestration of NRF2 by KEAP1 and suggest a possible regulatory step between KEAP1-NRF2 binding and NRF2 degradation.
- Novatchkova M, Bachmair A, Eisenhaber B, Eisenhaber F
- Proteins with two SUMO-like domains in chromatin-associated complexes: the RENi (Rad60-Esc2-NIP45) family.
- BMC Bioinformatics. 2005; 6: 22-22
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BACKGROUND: Post-translational modification by Small Ubiquitin-like Modifiers (SUMO) has been implicated in protein targeting, in the maintenance of genomic integrity and in transcriptional control. But the specific molecular effects of SUMO modification on many target proteins remain to be elucidated. Recent findings point at the importance of SUMO-mediated histone NAD-dependent deacetylase (HDAC) recruitment in transcriptional regulation. RESULTS: We describe the RENi family of SUMO-like domain proteins (SDP) with the unique feature of typically containing two carboxy-terminal SUMO-like domains. Using sequence analytic evidence, we collect family members from animals, fungi and plants, most prominent being yeast Rad60, Esc2 and mouse NIP45 http://mendel.imp.univie.ac.at/SEQUENCES/reni/. Different proteins of the novel family are known to interact directly with histone NAD-dependent deacetylases (HDACs), structural maintenance of chromosomes (SMC) proteins, and transcription factors. In particular, the highly non-trivial designation of the first of the two successive SUMO-domains in non-plant RENi provides a rationale for previously published functionally impaired mutant variants. CONCLUSIONS: Till now, SUMO-like proteins have been studied exclusively in the context of their covalent conjugation to target proteins. Here, we present the exciting possibility that SUMO domain proteins, similarly to ubiquitin modifiers, have also evolved in a second line - namely as multi-domain proteins that are non-covalently attached to their target proteins. We suggest that the SUMO stable fusion proteins of the RENi family, which we introduce in this work, might mimic SUMO and share its interaction motifs (in analogy to the way that ubiquitin-like domains mimic ubiquitin). This presumption is supported by parallels in the spectrum of modified or bound proteins e.g. transcription factors and chromatin-associated proteins and in the recruitment of HDAC-activity.
- Yoshitake Y et al.
- Proliferation potential-related protein, an ideal esophageal cancer antigen for immunotherapy, identified using complementary DNA microarray analysis.
- Clin Cancer Res. 2004; 10: 6437-48
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PURPOSE: To establish effective antitumor immunotherapy for esophageal cancer, we tried to identify an useful target antigen of esophageal cancer. EXPERIMENTAL DESIGN: We did cDNA microarray analysis to find a novel candidate antigen, proliferation potential-related protein (PP-RP). We examined cytotoxicity against tumor cells in vitro and in vivo of CTLs specific to PP-RP established from esophageal cancer patients. RESULTS: In 26 esophageal cancer tissues, an average of relative ratio of the expression of the PP-RP mRNA in cancer cells versus adjacent normal esophageal tissues was 396.2. Immunohistochemical analysis revealed that, in 20 of the 22 esophageal cancer tissues, PP-RP protein was strongly expressed only in the cancer cells and not so in normal esophageal epithelial cells. PP-RP protein contains 10 epitopes recognized by HLA-A24-restricted CTLs. These CTLs, generated from HLA-A24-positive esophageal cancer patients, had cytotoxic activity against cancer cell lines positive for both PP-RP and HLA-A24. Furthermore, adoptive transfer of the PP-RP-specific CTL line inhibited the growth of a human esophageal cancer cell line engrafted in nude mice. CONCLUSIONS: The expression of PP-RP in esophageal cancer cells was significantly higher than in normal cells, and the CTLs recognizing PP-RP killed tumor cells in vitro and also showed tumor rejection effects in a xenograft model. Therefore, PP-RP may prove to be an ideal tumor antigen useful for diagnosis and immunotherapy for patients with esophageal cancer. cDNA microarray analysis is a useful method to identify ideal tumor-associated antigens.
- Li T et al.
- Sumoylation of heterogeneous nuclear ribonucleoproteins, zinc finger proteins, and nuclear pore complex proteins: a proteomic analysis.
- Proc Natl Acad Sci U S A. 2004; 101: 8551-6
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SUMO, a small ubiquitin-related modifier, is known to covalently attach to a number of nuclear regulatory proteins such as p53, IkappaB, promyelocytic leukemia protein and c-Jun. The sumoylation reaction is catalyzed by the SUMO protease, which exposes the C-terminal active glycine residue of the nascent SUMO, the heterodimeric SUMO activating enzyme, the SUMO conjugating enzyme, Ubc9, and SUMO protein ligases, in a manner similar to ubiquitinylation. Identification of SUMO-regulated proteins is hampered by the fact that many sumoylated proteins are present at a level below normal detection limit. This limitation was overcome by either in vivo overexpression of Myc-SUMO or in vitro sumoylation with excess biotin-SUMO and Ubc9. Sumoylated proteins so obtained were affinity purified or isolated by immunoprecipitation. The isolated sumoylated proteins were identified by sequence analysis using mass spectrometric methods. Results reveal that several heterogeneous nuclear ribonucleoproteins (hnRNPs), zinc finger proteins, and nuclear pore complex proteins were sumoylated. The sumoylation of hnRNP A1, hnRNP F, and hnRNP K were confirmed in vivo by coimmunoprecipitation. In view of the facts that hnRNPs have been implicated in RNA splicing, transport, stability, and translation, our findings suggest that sumoylation could play an important role in regulating mRNA metabolism.
- Anantharaman V, Aravind L
- Novel conserved domains in proteins with predicted roles in eukaryotic cell-cycle regulation, decapping and RNA stability.
- BMC Genomics. 2004; 5: 45-45
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BACKGROUND: The emergence of eukaryotes was characterized by the expansion and diversification of several ancient RNA-binding domains and the apparent de novo innovation of new RNA-binding domains. The identification of these RNA-binding domains may throw light on the emergence of eukaryote-specific systems of RNA metabolism. RESULTS: Using sensitive sequence profile searches, homology-based fold recognition and sequence-structure superpositions, we identified novel, divergent versions of the Sm domain in the Scd6p family of proteins. This family of Sm-related domains shares certain features of conventional Sm domains, which are required for binding RNA, in addition to possessing some unique conserved features. We also show that these proteins contain a second previously uncharacterized C-terminal domain, termed the FDF domain (after a conserved sequence motif in this domain). The FDF domain is also found in the fungal Dcp3p-like and the animal FLJ22128-like proteins, where it fused to a C-terminal domain of the YjeF-N domain family. In addition to the FDF domains, the FLJ22128-like proteins contain yet another divergent version of the Sm domain at their extreme N-terminus. We show that the YjeF-N domains represent a novel version of the Rossmann fold that has acquired a set of catalytic residues and structural features that distinguish them from the conventional dehydrogenases. CONCLUSIONS: Several lines of contextual information suggest that the Scd6p family and the Dcp3p-like proteins are conserved components of the eukaryotic RNA metabolism system. We propose that the novel domains reported here, namely the divergent versions of the Sm domain and the FDF domain may mediate specific RNA-protein and protein-protein interactions in cytoplasmic ribonucleoprotein complexes. More specifically, the protein complexes containing Sm-like domains of the Scd6p family are predicted to regulate the stability of mRNA encoding proteins involved in cell cycle progression and vesicular assembly. The Dcp3p and FLJ22128 proteins may localize to the cytoplasmic processing bodies and possibly catalyze a specific processing step in the decapping pathway. The explosive diversification of Sm domains appears to have played a role in the emergence of several uniquely eukaryotic ribonucleoprotein complexes, including those involved in decapping and mRNA stability.
- Gao S, Scott RE
- Stable overexpression of specific segments of the P2P-R protein in human MCF-7 cells promotes camptothecin-induced apoptosis.
- J Cell Physiol. 2003; 197: 445-52
- Display abstract
The stable overexpression of near full-length P2P-R protein in human Saos 2 cells restricts cell cycle progression by inducing mitotic arrest at prometaphase and mitotic apoptosis (Gao and Scott, 2002). Those effects of P2P-R were observed in Saos-2 cells that lack p53 and employ a caspase-3-dependent apoptotic signaling pathway. The current studies were performed to evaluate if overexpression of specific segments of the P2P-R protein promote apoptosis in human MCF-7 cells that contain p53 and employ a different apoptotic signaling pathway. Since segments of P2P-R were found not to induce apoptosis independently, the ability of three different P2P-R segments to promote camptothecin-induced apoptosis was evaluated following their stable transfection and expression in MCF-7 cells. Relative to full-length P2P-R (1-1560 aa), the three P2P-R segments used in these studies included: P2P-R-2 (761-1560 aa), P2P-R-3 (1156-1560 aa), and P2P-R-4 (1314-1560 aa). The results document that overexpression of P2P-R-2 and P2P-R-3 promotes camptothecin-induced apoptosis by three to fivefold when assayed by flow cytometric analysis of apoptotic sub 2n cell populations or by TUNEL assays. In contrast, P2P-R-4 had no effect on apoptosis. These results suggest that the ability of P2P-R to promote camptothecin-induced apoptosis in MCF-7 cells involves a specific region (1156-1314 aa) that exists within P2P-R. The data presented also show that the p53 binding domain of P2P-R overlaps with the apoptosis-associated region and previous studies documented that this region of P2P-R also binds single-strand nucleotides (Witte and Scott, 1997). Therefore, P2P-R-promoted apoptosis induced by camptothecin may be influenced by such interactions.
- Haglund K, Di Fiore PP, Dikic I
- Distinct monoubiquitin signals in receptor endocytosis.
- Trends Biochem Sci. 2003; 28: 598-603
- Display abstract
Numerous cellular proteins are post-translationally modified by the addition of the small modifier protein ubiquitin (Ub). The functional consequences of the type of ubiquitination vary, such that polyubiquitinated proteins are targeted for degradation by the proteasome, whereas monoubiquitination is implicated in other cellular functions, including endocytic trafficking and DNA repair. The monoubiquitination of trafficking cargoes, such as receptors and associated proteins, as well as of endocytic accessory Ub-binding proteins, raises the question of the precise function of monoubiquitin signals in the endocytic route. Recent biochemical and genetic evidence shows that multiple monoubiquitination of epidermal growth factor and platelet-derived growth factor receptors provides trafficking and sorting tags that ensure receptor endocytosis and degradation, whereas monoubiquitination of accessory proteins might play a role in regulating their function as Ub-receptors in the endosome.
- Ohi MD, Vander Kooi CW, Rosenberg JA, Chazin WJ, Gould KL
- Structural insights into the U-box, a domain associated with multi-ubiquitination.
- Nat Struct Biol. 2003; 10: 250-5
- Display abstract
The structure of the U-box in the essential Saccharomyces cerevisiae pre-mRNA splicing factor Prp19p has been determined by NMR. The conserved zinc-binding sites supporting the cross-brace arrangement in RING-finger domains are replaced by hydrogen-bonding networks in the U-box. These hydrogen-bonding networks are necessary for the structural stabilization and activity of the U-box. A conservative Val-->Ile point mutation in the Prp19p U-box domain leads to pre-mRNA splicing defects in vivo. NMR analysis of this mutant shows that the substitution disrupts structural integrity of the U-box domain. Furthermore, comparison of the Prp19p U-box domain with known RING-E2 complex structures demonstrates that both U-box and RING-fingers contain a conserved interaction surface. Mutagenesis of residues at this interface, while not perturbing the structure of the U-box, abrogates Prp19p function in vivo. These comparative structural and functional analyses imply that the U-box and its associated ubiquitin ligase activity are critical for Prp19p function in vivo.
- Krishna SS, Majumdar I, Grishin NV
- Structural classification of zinc fingers: survey and summary.
- Nucleic Acids Res. 2003; 31: 532-50
- Display abstract
Zinc fingers are small protein domains in which zinc plays a structural role contributing to the stability of the domain. Zinc fingers are structurally diverse and are present among proteins that perform a broad range of functions in various cellular processes, such as replication and repair, transcription and translation, metabolism and signaling, cell proliferation and apoptosis. Zinc fingers typically function as interaction modules and bind to a wide variety of compounds, such as nucleic acids, proteins and small molecules. Here we present a comprehensive classification of zinc finger spatial structures. We find that each available zinc finger structure can be placed into one of eight fold groups that we define based on the structural properties in the vicinity of the zinc-binding site. Three of these fold groups comprise the majority of zinc fingers, namely, C2H2-like finger, treble clef finger and the zinc ribbon. Evolutionary relatedness of proteins within fold groups is not implied, but each group is divided into families of potential homologs. We compare our classification to existing groupings of zinc fingers and find that we define more encompassing fold groups, which bring together proteins whose similarities have previously remained unappreciated. We analyze functional properties of different zinc fingers and overlay them onto our classification. The classification helps in understanding the relationship between the structure, function and evolutionary history of these domains. The results are available as an online database of zinc finger structures.
- Li Z, Na X, Wang D, Schoen SR, Messing EM, Wu G
- Ubiquitination of a novel deubiquitinating enzyme requires direct binding to von Hippel-Lindau tumor suppressor protein.
- J Biol Chem. 2002; 277: 4656-62
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von Hippel-Lindau (VHL) disease is a hereditary cancer syndrome caused by germline mutations of the VHL gene. Recent studies suggest that VHL protein (pVHL) is a component of an E3 ubiquitin ligase, but the detailed biological function of pVHL remains to be determined. To further elucidate the biological functions of pVHL, we searched pVHL-interacting proteins using yeast two-hybrid screening. A novel protein named VHL-interacting deubiquitinating enzyme 1 (VDU1) was identified as being able to directly interact with pVHL in vitro and in vivo. We have determined the full-length cDNA of this enzyme, which includes two putative subtypes. Type I consists of 942 amino acids, and type II consists of 911 amino acids with predicted molecular masses of 107 and 103 kDa, respectively. We have also cloned a mouse homologue of this enzyme. Sequence analysis reveals that this protein is conserved between human and mouse and contains the signature motifs of the ubiquitin-specific processing protease family. Enzymatic function studies demonstrate its deubiquitinating activity. We have determined that the VDU1-interacting region in pVHL is located in its beta-domain, and several naturally occurring mutations located in this domain disrupt the interaction between pVHL and VDU1 protein. Co-immunoprecipitation demonstrates that VDU1 can be recruited into the pVHL-elongin C-elongin B complex. Finally, we demonstrate that VDU1 is able to be ubiquitinated via a pVHL-dependent pathway for proteasomal degradation, and VHL mutations that disrupt the interaction between VDU1 and pVHL abrogate the ubiquitination of VDU1. Our findings indicate that VDU1, a novel ubiquitin-specific processing protease, is a downstream target for ubiquitination and degradation by pVHL E3 ligase. Targeted degradation of VDU1 by pVHL could be crucial for regulating the ubiquitin-proteasome degradation pathway.
- Beitel LK et al.
- Cloning and characterization of an androgen receptor N-terminal-interacting protein with ubiquitin-protein ligase activity.
- J Mol Endocrinol. 2002; 29: 41-60
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The androgen receptor (AR) N-terminal domain plays a critical role in androgen-responsive gene regulation. A novel AR N-terminal-interacting protein (ARNIP) was isolated using the yeast two-hybrid system and its interaction with amino acids 11-172 of the normal or corresponding region of the polyglutamine-expanded human AR confirmed by glutathione S-transferase pulldown assays. ARNIP cDNAs cloned from NSC-34 (mouse neuroblastoma/spinal cord) or PC-3 (human prostate adenocarcinoma) mRNA encoded highly homologous 30 kDa (261 amino acids) cysteine-rich proteins with a RING-H2 (C3H2C3 zinc finger) domain; this motif is highly conserved in predicted ARNIP-homologous proteins from several other species. Expression of the approximately 1.7 kb ARNIP mRNA was detected in various tissues by Northern blotting, but was highest in mouse testes, kidney and several neuronal cell lines. In addition, the human ARNIP protein was found to be encoded by nine exons spanning 32 kb on chromosome 4q21. In COS-1 cells, coexpression of ARNIP and AR did not affect AR ligand-binding kinetics, nor did ARNIP act as a coactivator or corepressor in transactivation assays. However, AR N-terminal:C-terminal interaction was reduced in the presence of ARNIP. Intriguingly, ARNIP, and in particular its RING-H2 domain, functioned as a ubiquitin-protein ligase in vitro in the presence of a specific ubiquitin-conjugating enzyme, Ubc4-1. Mutation of a single cysteine residue in the ARNIP RING-H2 domain (Cys145Ala) abolished this E3 ubiquitin ligase activity. Fluorescent protein tagging studies revealed that AR-ARNIP interaction was hormone-independent in COS-1 cells, and suggest that colocalization of both AR and ARNIP to the nucleus upon androgen addition may allow ARNIP to play a role in nuclear processes. Thus, identification of a novel AR-interacting protein with ubiquitin ligase activity will stimulate further investigation into the role of ubiquitination and the ubiquitin-proteasome system in AR-mediated cellular functions.
- Gao S, Witte MM, Scott RE
- P2P-R protein localizes to the nucleolus of interphase cells and the periphery of chromosomes in mitotic cells which show maximum P2P-R immunoreactivity.
- J Cell Physiol. 2002; 191: 145-54
- Display abstract
P2P-R is a nuclear protein that can bind both p53 and Rb1. Its functions include roles in the control of RNA metabolism, apoptosis, and p53-dependent transcription. The expression of P2P-R also is repressed in G1 arrested terminally differentiated cells. The current studies therefore evaluated if P2P-R undergoes cell cycle-associated changes in its abundance and/or localization. Western blots show that relative to G0 quiescent cells, P2P-R protein levels are higher in populations of G2/M cells prepared by the physiological parasynchronization technique of serum deprivation followed by serum stimulation. More striking is the > 10-fold enrichment of P2P-R protein in specimens of highly purified mitotic cells prepared by the mitotic shake-select technique, or by synchrony with the mitotic spindle disruption agents nocodazole or vinblastine. These changes in P2P-R protein occur without a concomitant change in P2P-R mRNA expression suggesting that P2P-R immunoreactivity increases during mitosis. Confocal microscopy next established the localization of P2P-R to nucleoli in interphase cells and at the periphery of chromosomes in mitotic cells that lack nucleoli. The high levels of P2P-R localized to the periphery of chromosomes in mitotic cells suggest that P2P-R shares characteristics with other nucleolar proteins that associate with the periphery of chromosomes during mitosis. These include: nucleolin, B23, Ki67, and fibrillarin.
- Ardley HC, Tan NG, Rose SA, Markham AF, Robinson PA
- Features of the parkin/ariadne-like ubiquitin ligase, HHARI, that regulate its interaction with the ubiquitin-conjugating enzyme, Ubch7.
- J Biol Chem. 2001; 276: 19640-7
- Display abstract
We recently reported the identification of a RING finger-containing protein, HHARI (human homologue of Drosophila ariadne), which binds to the human ubiquitin-conjugating enzyme UbcH7 in vitro. We now demonstrate that HHARI interacts and co-localizes with UbcH7 in mammalian cells, particularly in the perinuclear region. We have further defined a minimal interaction region of HHARI comprising residues 186-254, identified individual amino acid residues essential for the interaction, and determined that the distance between the RING1 finger and IBR (in between RING fingers) domains is critical to maintaining binding. We have also established that the RING1 finger of HHARI cannot be substituted for by the highly homologous RING finger domains of either of the ubiquitin-protein ligase components c-CBL or Parkin, despite their similarity in structure and their independent capabilities to bind UbcH7. Furthermore, mutation of the RING1 finger domain of HHARI from a RING-HC to a RING-H2 type abolishes interaction with UbcH7. These studies demonstrate that very subtle changes to the domains that regulate recognition between highly conserved components of the ubiquitin pathway can dramatically affect their ability to interact.
- Katoh M
- Molecular cloning and characterization of RNF26 on human chromosome 11q23 region, encoding a novel RING finger protein with leucine zipper.
- Biochem Biophys Res Commun. 2001; 282: 1038-44
- Display abstract
Genetic alterations of RING finger genes, encoding an ubiquitin-protein ligase, are implicated in several types of human cancer through dysregulation of growth regulators. Here, a novel RING finger gene, RNF26, was cloned and characterized. The RNF26 gene on human chromosome 11q23 region was found to encode a polypeptide of 433 amino acids with the N-terminal leucine zipper domain and the C-terminal RING finger domain. Among the RING finger protein family, RING finger domains of RNF26, CGR19, NEURL, KIAA0554, and AK022937 were found to constitute a novel C3HC5 subfamily, which is distinct from C3H2C3 or C3HC4 subfamilies. RING finger domain of RNF26 was most homologous to that of CGR19 (49% amino-acid identity). The 3.2-kb RNF26 mRNA was expressed ubiquitously in normal human tissues, but was upregulated in several human cancer cell lines, including HL-60 (promyelocytic leukemia), HeLa S3 (cervical uterus cancer), SW480 (colorectal cancer), and MKN7 (gastric cancer). In addition, RNF26 was upregulated in 50% of primary gastric cancer examined in this study. Although substrates of ubiquitination mediated by RNF26 remain to be elucidated, RNF26 upregulation in several types of human cancer might be implicated in carcinogenesis through dysregulation of its substrates.
- Reed R, Magni K
- A new view of mRNA export: separating the wheat from the chaff.
- Nat Cell Biol. 2001; 3: 2014-2014
- Display abstract
Current models for the export of messenger RNA share the notion that the highly abundant class of nuclear RNA-binding proteins--the hnRNP proteins--have a key role in exporting RNA. But recent studies have led to a new understanding of several non-hnRNP proteins, including SR proteins and the conserved mRNA export factor ALY, which are recruited to the mRNA during pre-mRNA splicing. These studies, together with older work on hnRNP particles and assembly of the spliceosome, lead us to a new view of mRNA export. In our model, the non-hnRNP factors form a splicing-dependent mRNP complex that specifically targets mature mRNA for export, while hnRNP proteins retain introns in the nucleus. A machinery that is conserved between yeast and higher eukaryotes functions to export the mRNA.
- Joazeiro CA, Weissman AM
- RING finger proteins: mediators of ubiquitin ligase activity.
- Cell. 2000; 102: 549-52
- Aravind L, Koonin EV
- The U box is a modified RING finger - a common domain in ubiquitination.
- Curr Biol. 2000; 10: 1324-1324
- Jeanmougin F, Thompson JD, Gouy M, Higgins DG, Gibson TJ
- Multiple sequence alignment with Clustal X.
- Trends Biochem Sci. 1998; 23: 403-5
- Altschul SF et al.
- Gapped BLAST and PSI-BLAST: a new generation of protein database search programs.
- Nucleic Acids Res. 1997; 25: 3389-402
- Display abstract
The BLAST programs are widely used tools for searching protein and DNA databases for sequence similarities. For protein comparisons, a variety of definitional, algorithmic and statistical refinements described here permits the execution time of the BLAST programs to be decreased substantially while enhancing their sensitivity to weak similarities. A new criterion for triggering the extension of word hits, combined with a new heuristic for generating gapped alignments, yields a gapped BLAST program that runs at approximately three times the speed of the original. In addition, a method is introduced for automatically combining statistically significant alignments produced by BLAST into a position-specific score matrix, and searching the database using this matrix. The resulting Position-Specific Iterated BLAST (PSI-BLAST) program runs at approximately the same speed per iteration as gapped BLAST, but in many cases is much more sensitive to weak but biologically relevant sequence similarities. PSI-BLAST is used to uncover several new and interesting members of the BRCT superfamily.
- Hammarstrom A, Berndt KD, Sillard R, Adermann K, Otting G
- Solution structure of a naturally-occurring zinc-peptide complex demonstrates that the N-terminal zinc-binding module of the Lasp-1 LIM domain is an independent folding unit.
- Biochemistry. 1996; 35: 12723-32
- Display abstract
The three-dimensional solution structure of the 1:1 complex between the synthetic peptide ZF-1 and zinc was determined by 1H NMR spectroscopy. The peptide, initially isolated from pig intestines, is identical in sequence to the 30 N-terminal amino acid residues of the human protein Lasp-1 belonging to the LIM domain protein family. The final set of 20 energy-refined NMR conformers has an average rmsd relative to the mean structure of 0.55 A for the backbone atoms of residues 3-30. Calculations without zinc atom constraints unambiguously identified Cys 5, Cys 8, His 26, and Cys 29 as the zinc-coordinating residues. LIM domains consist of two sequential zinc-binding modules and the NMR structure of the ZF-1-zinc complex is the first example of a structure of an isolated module. Comparison with the known structures of the N-terminal zinc-binding modules of both the second LIM domain of chicken CRP and rat CRIP with which ZF-1 shares 50% and 43% sequence identity, respectively, supports the notion that the zinc-binding modules of the LIM domain have a conserved structural motif and identifies local regions of structural diversity. The similarities include conserved zinc-coordinating residues, a rubredoxin knuckle involving Cys 5 and Cys 8, and the coordination of the zinc ion by histidine N delta in contrast to the more usual coordination by N epsilon observed for other zinc-finger domains. The present structure determination of the ZF-1-zinc complex establishes the N-terminal half of a LIM domain as an independent folding unit. The structural similarities of N- and C-terminal zinc-binding modules of the LIM domains, despite limited sequence identity, lead to the proposal of a single zinc-binding motif in LIM domains. The coordinates are available from the Brookhaven protein data bank, entry 1ZFO.
- Frigerio JM et al.
- Analysis of 2166 clones from a human colorectal cancer cDNA library by partial sequencing.
- Hum Mol Genet. 1995; 4: 37-43
- Display abstract
Large scale sequencing of cDNAs from a tissue-specific library provides information on the functional phenotype of that tissue and the clones constitute a reservoir of biological markers. For these reasons, we have randomly sequenced 2166 clones of a cDNA library constructed with human colorectal cancer mRNAs. Database searches indicated that 1014 of the cDNAs represented known human genes or human homologs of other genes, 142 sequences corresponded to known ESTs, 119 sequences corresponded to 28S rRNA, repetitive sequences or poly(A) stretches only, and 891 corresponded to unknown transcripts representing the products of 740 new genes. Preliminary studies demonstrated that expression of some of them was altered in cancer. That cDNA collection is therefore a source of potential markers of colorectal cancer.