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103 records returned

Kinetic and chemical characterization of aldehyde oxidation by fungal aryl-alcohol oxidaseEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Fungal aryl-alcohol oxidase (AAO) provides H2O2 for lignin biodegradation. AAO is active on benzyl alcohols that are oxidized to aldehydes. However, the H2O2 formed from some of them was more than stoichiometric with respect to the aldehyde detected. This was due to a double reaction that involves aryl-aldehyde oxidase activity. The latter was investigated using different benzylic aldehydes, whose oxidation to acids was demonstrated by GC-MS. The steady and pre-steady state kinetic constants together with the chromatographic results revealed a strong influence of substrate electron withdrawing/donating substituents on acti...
Source: BJ Structure - November 5, 2009 Category: Biochemistry Authors: P Ferreira, A Hernández-Ortega, B Herguedas, J Rencoret, A Gutiérrez, M Martínez, J Jiménez-Barbero, M Medina, Á T. Martínez Tags: BJ Structure Source Type: journals

Structural insights into the catalytic mechanism of Trypanosoma cruzi GPXI {-} glutathione peroxidase-like enzyme IEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Current drug therapies against Trypanosoma cruzi, causative agent of Chagas disease, have limited effectiveness and are highly toxic. T. cruzi-specific metabolic pathways that utilise trypanothione for the reduction of peroxides are being explored as potential novel therapeutic targets. We solved the X-ray crystal structure of one of the T. cruzi enzymes involved in peroxide reduction, the glutathione peroxidase-like enzyme TcGPXI. We also characterized the wild-type, Cys48Gly and Cys96Gly variants of TcGPXI by NMR spectroscopy and biochemical assays. Our data show that residues Cys48 and Cys96 are required for catalytic a...
Source: BJ Structure - November 3, 2009 Category: Biochemistry Authors: S Patel, S Hussain, R Harris, S Sardiwal, J M Kelly, S R Wilkinson, P C Driscoll, S Djordjevic Tags: BJ Structure Source Type: journals

Chicken ileal bile acid binding protein: a promising target of investigation to understand binding cooperativity across the protein familyEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Protein-bile acid interactions are crucial microscopic events at the basis of both physiological and pathological biochemical pathways. Bile acid binding proteins are intracellular transporters able to bind ligands with different stoichiometry, selectivity, and cooperativity. The molecular determinants and energetics of interaction are the observables that connect the microscopic to the macroscopic frameworks. This paper addresses the study and proposes a mechanism for the multi-site interaction of bile acids with chicken ileal bile acid binding protein (I-BABP) with the aim of elucidating the determinants of ligand bindin...
Source: BJ Structure - October 29, 2009 Category: Biochemistry Authors: M Guariento, M Assfalg, S Zanzoni, D Fessas, R Longhi, H Molinari Tags: BJ Structure Source Type: journals

Crystal structure of the human transcription elongation factor DSIF hSpt4 subunit in complex with the hSpt5 dimerization interfaceEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
The eukaryotic transcription elongation factor 5,6-dichloro-1-β-D-ribofuranosylbenzimidazole (DRB) sensitivity inducing factor (DSIF) is composed of two subunits, hSpt4 and hSpt5, which are homologous to the yeast factors Spt4 and Spt5. DSIF is involved in regulating the processivity of RNA polymerase II and plays an essential role in transcriptional activation of eukaryotes. At several eukaryotic promoters, DSIF together with the negative elongation factor NELF, leads to promoter-proximal pausing of RNA polymerase II. Here we describe the crystal structure of hSpt4 in complex with the dimerization region of hSpt5 (...
Source: BJ Structure - October 27, 2009 Category: Biochemistry Authors: S Wenzel, B M. Martins, P Rösch, B M. Wöhrl Tags: BJ Structure Source Type: journals

The oxyanion hole of Pseudomonas fluorescens mannitol 2-dehydrogenase: a novel structural motif for electrostatic stabilisation in alcohol dehydrogenase active sitesEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
The side chains of Asn-191 and Asn-300 constitute a characteristic structural motif of the active site of Pseudomonas fluorescens mannitol 2-dehydrogenase that lacks precedent in known alcohol dehydrogenases and resembles the canonical oxyanion binding pocket of serine proteases. We have used steady-state and transient kinetic studies of the effects of varied pH and deuterium isotopic substitutions in substrates and solvent on the enzymatic rates to delineate catalytic consequences resulting from individual and combined replacements of the two asparagines by Ala. The rate constants for the overall hydride transfer to and f...
Source: BJ Structure - October 26, 2009 Category: Biochemistry Authors: M Klimacek, B Nidetzky Tags: BJ Structure Source Type: journals

The molecular basis of the effect of temperature on enzyme activityEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Experimental data show that the effect of temperature on enzymes cannot be adequately explained in terms of a two-state model based on increases in activity and denaturation. The Equilibrium Model provides a quantitative explanation of enzyme thermal behaviour under reaction conditions by introducing an inactive (but not denatured) intermediate in rapid equilibrium with the active form. The temperature mid-point (Teq) of the rapid equilibration between the two forms is related to the growth temperature of the organism, and the enthalpy of the equilibrium (∆Heq) to its ability to function over various temperature ran...
Source: BJ Structure - October 21, 2009 Category: Biochemistry Authors: R M. Daniel, M E. Peterson, M J. Danson, N C. Price, S M. Kelly, C R. Monk, C S. Weinberg, M L. Oudshoorn, C K. Lee Tags: BJ Structure Source Type: journals

An allosteric transition trapped in an intermediate state of a new kinesin-inhibitor complexEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Human kinesin Eg5 plays an essential role in mitosis by separating duplicated centrosomes and establishing the bipolar spindle. Eg5 is an interesting drug target for the development of cancer chemotherapy, with 7 inhibitors already in clinical trials. Here, we report the crystal structure of the Eg5 motor domain complexed with a potent antimitotic inhibitor S-trityl-L-cysteine to 2.0 Å resolution. The Eg5-STLC complex crystallises in space group P32 with 3 molecules per asymmetric unit. Two of the molecules reveal the final inhibitor-bound state of Eg5, whereby Loop L5 has swung downwards to close the inhibitor-bind...
Source: BJ Structure - September 29, 2009 Category: Biochemistry Authors: H Kaan, V Ulaganathan, D D. Hackney, F Kozielski Tags: BJ Structure Source Type: journals

Structural insight on the control of urea synthesis: identification of the binding site for N-acetyl-L-glutamate, the essential allosteric activator of mitochondrial carbamoyl phosphate synthetaseEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
N-acetyl-L-glutamate (NAG), the essential allosteric activator of the first urea cycle enzyme, carbamoyl-phosphate synthetase (CPSI), is a key regulator of this crucial cycle for ammonia-detoxification in animals (including humans). Automated cavity searching and flexible docking have allowed identification of the NAG site in the crystal structure of human CPSI C-terminal domain. The site, a pocket lined by invariant residues and located between the central β-sheet and two α-helices, opens at the β-sheet C-edge and is roofed by a 3-residue lid. It can tightly accommodate one extended NAG molecule havin...
Source: BJ Structure - September 14, 2009 Category: Biochemistry Authors: S Pekkala, A Isabel Martínez, B Barcelona, J Gallego, E Bendala, I Yefimenko, V Rubio, J Cervera Tags: BJ Structure Source Type: journals

Mechanistic insights into Cu(I) cluster transfer between the chaperone CopZ and its cognate Cu(I)-transporting P-type ATPase, CopAEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Multinuclear Cu(I) clusters are common in nature, but little is known about their formation or transfer between proteins. CopZ and CopA from Bacillus subtilis, which are involved in a copper-efflux pathway, both readily accommodate multinuclear Cu(I) clusters. Using the luminescence properties of a multinuclear Cu(I)-bound form of the two N-terminal soluble domains of CopA (CopAab) we have investigated thermodynamic and kinetic properties of cluster formation and loss. We demonstrate that Cu(I)-bound forms of dimeric CopZ containing more than 1 Cu(I) per CopZ monomer can transfer Cu(I) to apo-CopAab leading to the formatio...
Source: BJ Structure - September 13, 2009 Category: Biochemistry Authors: C Singleton, S Hearnshaw, L Zhou, N E Le Brun, A M Hemmings Tags: BJ Structure Source Type: journals

A survey of proteins encoded by non-synonymous SNPs reveals a significant fraction with altered stability and activityEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
On average, each human gene has about four single nucleotide polymorphisms (SNPs) in the coding region, half of which are non-synonymous or missense SNPs (nsSNP). Current attention is focused on those that are known to perturb function and are strongly linked to disease. However, the vast majority of SNPs have not been investigated for the possibility of causing disease. We set out to assess the fraction of nsSNPs that encode proteins that have altered stability and activity, for this class of variants would be candidates to perturb cellular function. We tested the thermostability and, where possible, the catalytic activit...
Source: BJ Structure - August 24, 2009 Category: Biochemistry Authors: A Allali-Hassani, G A. Wasney, I Chau, B Hong, G Senisterra, P Loppnau, Z Shi, J Moult, A M. Edwards, C H. Arrowsmith, H Park, M Schapira, M Vedadi Tags: BJ Structure Source Type: journals

Fluorine substitutions in an antigenic peptide selectively modulate T cell receptor binding in a minimally perturbing mannerEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
T cell receptor (TCR) recognition of antigenic peptides bound and presented by major histocompatibility complex (MHC) molecules forms the basis of the cellular immune response to pathogens and cancer. TCRs bind peptide/MHC molecules weakly and with fast kinetics, features which have hindered detailed biophysical studies of these interactions. Modified peptides resulting in enhanced TCR binding could help overcome these challenges. Further, there is considerable interest in using modified peptides with enhanced TCR binding as the basis for clinical vaccines. Here, we studied how fluorine substitutions in an antigenic peptid...
Source: BJ Structure - August 20, 2009 Category: Biochemistry Authors: K H. Piepenbrink, O Y. Borbulevych, R F. Sommese, J Clemens, K M Armstrong, C Desmond, P Do, B M. Baker Tags: BJ Structure Source Type: journals

The crystal structure of caspase-6, a selective effector of axonal degeneration.Email this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
We report here biochemical and structural studies on caspase-6. As an effector caspase, caspase-6 is a constitutive dimer independent of the maturation state of the enzyme. The ligand-free structure shows caspase-6 in a partially mature but latent conformation. The cleaved inter-domain linker remains partially inserted in the central groove of the dimer as observed in other caspases. However, in contrast to previous structures, not only is the catalytic machinery mis-aligned, but several structural elements required for substrate recognition are missing. Most importantly, residues forming a short anti-parallel β-she...
Source: BJ Structure - August 20, 2009 Category: Biochemistry Authors: R Baumgartner, G Meder, C Briand, A Decock, A D'Arcy, U Hassiepen, R Morse, M Renatus Tags: BJ Structure Source Type: journals

C-terminal sequences of hsp70 and hsp90 as non-specific anchors for TPR proteinsEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Synopsis Steroid hormone receptor maturation is a multi-step process that involves several TPR proteins that bind to the maturation complex via the C-termini of hsp70 and hsp90. We produced a random T7 peptide library to investigate the roles played by the C-termini of the two heat shock proteins in the TPR/hsp interactions. Surprisingly, phages with the MEEVD sequence, found at the C-terminus of hsp90, were not recovered from our biopanning experiments. However, two groups of phages were isolated that bound relatively tightly to HsPP5 TPR. Multiple copies of phages with a C-terminal sequence of LFG were isolated. These...
Source: BJ Structure - August 17, 2009 Category: Biochemistry Authors: A John Ramsey, L C Russell, M Chinkers Tags: BJ Signal Source Type: journals

The carboxy-terminal domain is sufficient for endonuclease activity of Neisseria gonorrhoeae MutLEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
We report here that the C-terminal domain of Neisseria gonorrhoeae MutL (NgoL-CTD) consisting of amino acids 460 to 658 exhibits Mn2+ dependent endonuclease activity. Sedimentation velocity,sedimentation equilibrium and dynamic light scattering experiments show NgoL-CTD to be a dimer. The probable endonucleolytic active site is localized to a metal binding motif, DMHA(X)2E(X) 4E and the nicking endonuclease activity is dependent on the integrity of this motif. By in vitro comparison of wild-type and a mutant NgoL-CTD protein, we show that the latter protein exhibits highly reduced endonuclease activity. We therefore...
Source: BJ Structure - August 5, 2009 Category: Biochemistry Authors: V Duppatla, C Bodda, C Urbanke, P Friedhoff, D N Rao Tags: BJ Gene Source Type: journals

Structure of a trimeric bacterial microcompartment shell protein, EtuB, associated with ethanol utilisation in Clostridium kluyveriEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
It has been suggested that ethanol metabolism in the strict anaerobe Clostridium kluyveri occurs within a metabolosome, a subcellular proteinaceous bacterial microcompartment. Two bacterial microcompartment shell proteins (EtuA and EtuB) are found encoded on the genome clustered with the genes for ethanol utilisation. The function of the bacterial microcompartment is to facilitate fermentation by sequestering the enzymes, substrates, and intermediates. Recent structural studies of bacterial microcompartment proteins have revealed both hexamers and pentamers that assemble to generate the pseudo-icosahedral bacterial microco...
Source: BJ Structure - July 27, 2009 Category: Biochemistry Authors: D Heldt, S Frank, A Seyedarabi, D Ladakis, J B Parsons, M J Warren, R W Pickersgill Tags: BJ Structure Source Type: journals

Binding of the CHD4 PHD2 finger to histone H3 is modulated by covalent modificationsEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Chromodomain helicase DNA-binding protein 4 (CHD4) ATPase is a major subunit of the repressive NuRD (nucleosome remodeling and deacetylase) complex, which is involved in transcriptional regulation and development. CHD4 contains two plant homeodomain (PHD) fingers of unknown function. Here we show that the second PHD finger (PHD2) of CHD4 recognizes the amino-terminus of histone H3 and that this interaction is facilitated by acetylation or methylation of Lys9 (H3K9ac and H3K9me, respectively) but is inhibited by methylation of Lys4 (H3K4me) or acetylation of Ala1 (H3A1ac). An 18 μM binding affinity toward unmodified ...
Source: BJ Structure - July 21, 2009 Category: Biochemistry Authors: C A Musselman, R E Mansfield, A L Garske, F Davrazou, A H Kwan, S S Oliver, H OLeary, J M Denu, J P Mackay, T G Kutateladze Tags: BJ Structure Source Type: journals

Development of Dictyostelium discoideum is associated with alteration of fucosylated N-glycan structuresEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
The social amoeba Dictyostelium discoideum has become established as a simple model for the examination of cell-cell interactions and early studies suggested that shifts in glycosylation profiles take place during its life cycle. In the present study, we have applied HPLC and mass spectrometric methods to show that the major N-glycans in axenic cultures of the AX3 strain are oligomannosidic forms, most of which carry core fucose and/or intersecting and bisecting N-acetylglucosamine residues, including the major structure with the composition Man8GlcNAc4Fuc1. The postulated α1,3-linkage of the core fucose which corre...
Source: BJ Structure - July 16, 2009 Category: Biochemistry Authors: B Schiller, A Hykollari, J Voglmeir, G Pöltl, K Hummel, E Razzazi-Fazeli, R Geyer, I B. H. Wilson Tags: BJ Structure Source Type: journals

Mapping of the ligand binding site on the b{'} domain of human PDI; interaction with peptide ligands and the x-linker regionEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
In this study we show that the b’ domain of human PDI is in conformational exchange but that its structure is stabilised by the addition of peptide ligands or by binding the x-linker region. The location of the ligand binding site in b’ was mapped by NMR chemical shift perturbation and found to consist primarily of residues from the core β-sheet and helices 1 and 3. This site is where the x-linker region binds in the x-ray structure of b’x and we show that peptide ligands can compete with x binding at this site. The finding that x binds in the principal ligand binding site of b’ further s...
Source: BJ Structure - July 14, 2009 Category: Biochemistry Authors: L J. Byrne, A Sidhu, A Katrine Wallis, L W. Ruddock, R B. Freedman, M J. Howard, R A. Williamson Tags: BJ Structure Source Type: journals

Structural basis of transport of lysophospholipids by human serum albuminEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
This study provides structural information on HSA-lysophospholipids interaction and may facilitate the understanding of the transport and distribution of lysophospholipids. (Source: BJ Structure)
Source: BJ Structure - July 13, 2009 Category: Biochemistry Authors: S Guo, X Shi, F Yang, L Chen, E J. Meehan, C Bian, M Huang Tags: BJ Structure Source Type: journals

Pup, a prokaryotic ubiquitin-like protein, is an intrinsically disordered proteinEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Pup from Mycobacterium tuberculosis (Mtb) is the first ubiquitin-like protein identified in non-eukaryotic cells. Though different ubiquitin-like proteins from eukaryotes share low sequence similarity, their three-dimensional (3D) structures exhibit highly conserved typical ubiquitin-like fold. Interestingly, our studies reveal that Pup not only shares low sequence similarity but also presents a totally distinguished structure compared with other ubiquitin-like superfamily proteins. Diverse structure predictions combined with CD and NMR spectroscopic studies all demonstrate that Pup is an intrinsically disordered protein. ...
Source: BJ Structure - July 6, 2009 Category: Biochemistry Authors: S Liao, Q Shang, X Zhang, J Zhang, C Xu, X Tu Tags: BJ Structure Source Type: journals

Molecular recognition of physiological substrate noradrenaline by the adrenaline synthesising enzyme PNMT and factors influencing its methyltransferase activityEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Substrate specificity is critically important for enzyme catalysis. In the adrenaline synthesising enzyme phenylethanolamine N-methyltransferase (PNMT), minor changes in substituents can convert substrates into inhibitors. Here we report the crystal structures of 6 human PNMT complexes including the first structure of the enzyme in complex with its physiological ligand R-noradrenaline: determining this structure required rapid soak methods because of the tendency for noradrenaline to oxidize. Comparison of the PNMT:noradrenaline complex with the previously determined PNMT:octopamine complex demonstrates that these two subs...
Source: BJ Structure - July 1, 2009 Category: Biochemistry Authors: N Drinkwater, C L Gee, M Puri, K R Criscione, M J McLeish, G L Grunewald, J L Martin Tags: BJ Structure Source Type: journals

The transcriptional activity of Pygopus is enhanced by its interaction with CREB binding proteinEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
In this report, we demonstrate that Pygopus can interact with the histone acetyltransferase (HAT) cyclic AMP responsive element binding protein (CREB) binding protein (CBP). The interaction is via the N-terminal homology domain (NHD) of Pygopus which binds to two regions in the HAT domain of CBP. Transfected and endogenous hPygo2 and CBP proteins co-immunoprecipitate in human embryonic kidney (HEK293) cells and both proteins co-localize in SW480 colorectal cancer cells. The interaction with CBP also enhances both DNA-tethered and TCF/LEF1-dependent transcriptional activity of Pygopus. Furthermore, immunoprecipitated Pygopu...
Source: BJ Structure - June 24, 2009 Category: Biochemistry Authors: P G.P. Andrews, Z He, C Popadiuk, K R. Kao Tags: BJ Structure Source Type: journals

Crystal structure of Glycine max glutathione transferase in complex with glutathione: investigation of the mechanism operating by the tau class glutathione transferasesEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Cytosolic glutathione transferases (GSTs) are a multifunctional group of enzymes widely distributed in nature and involved in cellular detoxification processes. The three-dimensional structure of Glycine max GSTU4-4 (GmGSTU4-4) complexed with glutathione (GSH) was determined by the molecular replacement method at 2.7 Å resolution. The bound GSH is located in a region formed by the beginning of α-helices H1, H2 and H3 in the N-terminal domain of the enzyme. Significant differences in the GSH binding site (G-site) as compared to the structure determined in complex with S-(p-nitrobenzyl)-glutathione (Nb-GSH) wer...
Source: BJ Structure - June 18, 2009 Category: Biochemistry Authors: I Axarli, P Dhavala, A C. Papageorgiou, N E Labrou Tags: BJ Structure Source Type: journals

Gain of function mutations identify amino acids within transmembrane domains of the yeast vacuolar transporter Zrc1 that determine metal specificityEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Cation diffusion facilitator transporters are found in all three kingdoms of life and are involved in transporting transition metals out of the cytosol. The metals they transport include: Zn2+, Co2+, Fe2+, Cd2+, Ni2+, Mn2+, however, no single transporter transports all metals. Previously we showed that a single amino acid mutation in the yeast vacuolar zinc transporter Zrc1 changed its substrate specificity from Zn2+ to Fe2+ and Mn2+ (Lin et al, (2008) J. Biol. Chem. 283: 33865-33873). Mutant Zrc1 that gained iron transport activity could protect cells with a d...
Source: BJ Structure - June 17, 2009 Category: Biochemistry Authors: H Lin, D Burton, L Li, D E Warner, J Phillips, D McVey Ward, J Kaplan Tags: BJ Metabolism Source Type: journals

Specific inhibition of tissue kallikrein 1 with a human monoclonal antibody reveals potential role in airway diseasesEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Tissue kallikrein 1 (KLK1) is a member of the tissue kallikrein family of serine proteases and the primary kinin generating enzyme in human airways. DX-2300 is a fully human antibody that inhibits KLK1 via a competitive inhibition mechanism (Ki = 0.13 nM). No binding of DX-2300 to KLK1 was observed in a surface plasmon resonance biosensor assay when KLK1 was complexed to known active site inhibitors, suggesting that DX-2300 recognizes the KLK1 active site. DX-2300 did not inhibit any of the 21 serine proteases that were tested at concentration of 1 µM. We validated the use of DX-2300 for specific KLK1 inhibition by ...
Source: BJ Structure - June 15, 2009 Category: Biochemistry Authors: D J Sexton, T Chen, D Martik, P Kuzmic, G Kuang, J Chen, A E Nixon, B L Zuraw, R M Forteza, W M. Abraham, C R. Wood Tags: BJ Disease Source Type: journals

Modulation of the proteolytic activity of the complement protease C1s by polyanions: implications for polyanion-mediated acceleration of interaction between C1s and SERPING1Email this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Complement plays crucial roles in the immune system, but incorrect regulation causes inflammation and targeting of self-tissue, leading to diseases such as systemic lupus erythematosus, rheumatoid arthritis and age-related macular degeneration. In vivo, the initiating complexes of the classical and lectin pathways are controlled by SERPING1 (C1-inhibitor), which inactivates their C1s and MASP-2 protease components. Glycosaminoglycans (GAGs) and dextran sulfate (DXS) are able to significantly accelerate SERPING1-mediated inactivation of the key effector enzyme of the classical C1 complex, C1s, though the mechanism is poorly...
Source: BJ Structure - June 11, 2009 Category: Biochemistry Authors: T A. Murray-Rust, F K. Kerr, A René Thomas, T Wu, Y Tang, P Ong, N S Quinsey, J C Whisstock, I G.A. Wagenaar-Bos, C Freeman, R Neil Pike Tags: BJ Structure Source Type: journals

Differential contributions of Glu96, Asp102 and Asp111 to coagulation factor V/Va metal ion binding and subunit stabilityEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
This study defines the highly conserved segment spanning Glu96-Asp111 in FV as multifunctional. Of the three amino acids evaluated, Asp111 is essential and likely functions through direct and indirect effects on Ca2+ and Cu2+ interactions. Glu96 and Asp102 individually influence FV/FVa by more subtle effects possibly at the metal ion-dependent subunit interface. (Source: BJ Structure)
Source: BJ Structure - June 11, 2009 Category: Biochemistry Authors: J Song, K Talbot, J Hewitt, R T.A. MacGillivray, E L.G. Pryzdial Tags: BJ Structure Source Type: journals

The 5' terminal uracil of let-7a is critical for the recruitment of mRNA to Argonaute2Email this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Small RNAs modulate gene expression by forming a ribonucleoprotein complex with Argonaute proteins and direct them to specific complementary sites in target nucleic acids. However, the interactions required for the recruitment of target nucleic acid to the ribonucleoprotein complex are poorly understood. Here, we have investigated this question by using let-7a, Argonaute2 and a fully complementary mRNA target. Importantly, we have found recombinant Argonaute2 is sufficient to direct let-7a guided cleavage of mRNA. Thus this model system has allowed us to investigate the mechanistic basis of silencing in vitro and in vivo. ...
Source: BJ Structure - June 9, 2009 Category: Biochemistry Authors: K M Felice, D W Salzman, J Shubert-Coleman, K P Jensen, H M Furneaux Tags: BJ Structure Source Type: journals

Crystal structure of an inverting GH43 1,5-{alpha}-L-arabinanase from Geobacillus stearothermophilus complexed with its substrateEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Arabinanases are glycosidases that hydrolyze alpha-(1→5)- arabinofuranosidic linkages found in the backbone of the pectic polysaccharide arabinan. Here we describe the biochemical characterization and the enzyme-substrate crystal structure of an inverting family 43 arabinanase, from Geobacillus stearothermophilus T-6 (AbnB). Based on viscosity and reducing power measurements, and based on products analysis for the hydrolysis of linear arabinan by AbnB, the enzyme work in an endo mode of action. Isothermal titration calorimetry studies of a catalytic mutant with various arabinooligosaccharides suggested that the enzy...
Source: BJ Structure - June 8, 2009 Category: Biochemistry Authors: A Alhasid, A Ben-David, O Tabachnikov, D Wolf, E Naveh, G Zolotnitsky, Y Shoham, G Shoham Tags: BJ Structure Source Type: journals

The RNAP subunits F/E (RPB4/7) are stably associated with archaeal RNA polymerase - using fluorescence anisotropy to monitor RNAP assembly in vitroEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Archaeal and eukaryotic RNAPs are comprised of twelve subunits with discrete functions. Two of the subunits, F/E, have been hypothesised to associate with RNAP in a reversible manner during the transcription cycle. We have characterised the molecular interactions between F/E and the RNAP core. F/E binds to RNAP with submicromolar affinity but is not in a dynamic exchange with unbound F/E. (Source: BJ Structure)
Source: BJ Structure - June 3, 2009 Category: Biochemistry Authors: D Grohmann, A Hirtreiter, F Werner Tags: BJ Gene Source Type: journals

Helicobacter pylori UreE, a urease accessory protein: specific Ni2{+} and Zn2{+} binding properties and interaction with its cognate UreGEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
This study focuses on the metal-binding properties of UreE from Helicobacter pylori (HpUreE) and its interaction with the related accessory protein HpUreG, a GTPase involved in the assembly of the urease active site. Isothermal titration calorimetry (ITC) showed that HpUreE binds one equivalent of Ni2+ (Kd = 0.15 mM) or Zn2+ (Kd = 0.49 mM) per dimer, without modification of the protein oligomeric state, as indicated by light scattering. Different ligand environments for Zn2+ and Ni2+, which involve crucial histidine residues, were revealed by site-directed mutagenesis, suggesting a mechanism for...
Source: BJ Structure - May 29, 2009 Category: Biochemistry Authors: M Bellucci, B Zambelli, F Musiani, P Turano, S Ciurli Tags: BJ Structure Source Type: journals

FAM20B is a kinase that phosphorylates xylose in the glycosaminoglycan-protein linkage regionEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
In this study, we identified FAM20B as a kinase that phosphorylates the xylose residue in the linkage region. Overexpression of fam20b increased the amount of both chondroitin sulfate and heparan sulfate in HeLa cells, while the RNA interference of fam20b resulted in a reduction of their amount in the cells. Gel filtration analysis of the glycosaminoglycan chains synthesized in the overexpressing cells revealed that the glycosaminoglycan chains had similar length to those in mock-transfected cells. These results suggest that FAM20B regulates the number of glycosaminoglycan chains by phosphorylating the xylose residue in th...
Source: BJ Structure - May 28, 2009 Category: Biochemistry Authors: T Koike, T Izumikawa, J Tamura, H Kitagawa Tags: BJ BJ Structure Source Type: journals

Copper(I)-mediated protein-protein interactions result from suboptimal interaction surfacesEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
The homeostasis of metal ions in cells is the result of the contribution of several cellular pathways that involve transient, often weak, protein-protein interactions. Metal transfer typically implies the formation of adducts where the metal itself acts as a bridge between proteins, by coordinating residues of both interacting partners. Here we addressed the interaction between the human copper(I)-chaperone HAH1 and one metal-binding domain of one of its partners, namely the P-type copper-transporting ATPase ATP7A. The adduct was structurally characterized in solution, in the presence of copper(I), and through X-ray crysta...
Source: BJ Structure - May 19, 2009 Category: Biochemistry Authors: L Banci, I Bertini, V Calderone, N Della-Malva, I Caterina Felli, S Neri, A Pavelkova, A Rosato Tags: BJ BJ Structure Source Type: journals

Artificial {beta}-defensin based on a minimal defensin templateEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
We have designed and chemically synthesised an artificial β-defensin based on a minimal template derived from the comparative analysis of over 80 naturally occurring sequences. This molecule has the disulfide-bridged, β-sheet core structure of natural β-defensins and shows a robust, salt-sensitive antimicrobial activity against bacteria and yeast, as well as a chemotactic activity against immature dendritic cells. An SAR study using two truncated fragments or a Cys→Ser point-mutated analogue, in which one or two of the three disulphide bridges were absent, indicated that altering the structure r...
Source: BJ Structure - May 19, 2009 Category: Biochemistry Authors: N Antcheva, F Morgera, L Creatti, L Vaccari, U Pag, S Pacor, Y Shai, H Sahl, A Tossi Tags: BJ BJ Structure Source Type: journals

Bactericidal and membrane disruption activities of the eosinophil cationic protein are largely retained in an N-terminal fragmentEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Eosinophil cationic protein (ECP) is an eosinophil secretion protein with antipathogen activities involved in the host immune defense system. The bactericidal capacity of ECP relies on its action on both the plasma membrane and the bacterial wall. In a search for the structural determinants of ECP antimicrobial activity, we have identified an N-terminal domain (residues 1-45) that retains most of ECP’s membrane-destabilizing and antimicrobial activities. Two sections of this domain, ECP(1-19) and ECP(24-45), have also been evaluated. All three peptides bind and partially insert into lipid bilayers, inducing aggregat...
Source: BJ Structure - May 18, 2009 Category: Biochemistry Authors: M Torrent, B G De la Torre, M Nogués, D Andreu, E Boix Tags: BJ BJ Cell Source Type: journals

Variably modulated gating of the 26S proteasome by ATP and polyubiquitinEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
The 26S proteasome is a 2,500,000-dalton protease complex that degrades polyubiquitylated proteins by a mechanism that requires ATP hydrolysis. It also degrades short non-ubiquitylated peptides and certain unstructured proteins by an energy-independent mechanism that requires bound ATP to maintain its component subcomplexes, the 20S proteasome and PA700, in a functionally assembled state. Proteolysis of both types of substrates requires PA700-induced opening of reversible gates at substrate access pores of the 20S proteasome. Here we demonstrate that the rate of peptide substrate hydrolysis, a functional monitor of gate o...
Source: BJ Structure - May 13, 2009 Category: Biochemistry Authors: X Li, G N. DeMartino Tags: BJ BJ Cell Source Type: journals

Anti-parallel {beta}-sheet - a signature structure of the oligomeric amyloid-beta peptideEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Alzheimer’s disease (AD) is linked to amyloid beta peptide (Aβ) misfolding. Studies demonstrate that the level of soluble Aβ oligomeric forms correlates better with the progression of the disease than the level of fibrillar forms. Conformation-dependent antibodies have been developed to detect either Aβ oligomers or fibrils, suggesting that structural differences between these forms of Aβ exist. Using conditions which yield well-defined Aβ(1-42) oligomers or fibrils, we studied the secondary structure of these species by attenuated total reflection-FTIR spectroscopy. Whereas fibril...
Source: BJ Structure - May 13, 2009 Category: Biochemistry Authors: E Cerf, R Sarroukh, S Tamamizu-Kato, L Breydo, S Derclaye, Y Dufrêne, V Narayanaswami, E Goormaghtigh, J Ruysschaert, V Raussens Tags: BJ BJ Structure Source Type: journals

Carbohydrate binding domain on galectin-1 is more extensive for a complex glycan than for simple saccharides: implications for galectin-glycan interactions at the cell surfaceEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Galectin-1 (gal-1) mediates cell-cell and cell-extracellular matrix adhesion, essentially by interacting with β-galactoside-containing glycans of cell surface glycoconjugates. While most structural studies with gal-1 have investigated its binding to simple carbohydrates, in particular lactose and N-acetyllactosamine, this view is limited because gal-1 functions at the cell surface by interacting with more complex glycans that are heterogeneous in size and composition. Here, we used NMR spectroscopy to investigate the interaction of human gal-1 to a large (120 kDa), complex glycan [GRG: α-(1→2)-L-rhamno...
Source: BJ Structure - May 12, 2009 Category: Biochemistry Authors: M Miller, I V Nesmelova, D Platt, A Klyosov, K H Mayo Tags: BJ BJ Structure Source Type: journals

Quinalizarin as a potent, selective and cell permeable inhibitor of protein kinase CK2Email this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Emodin (1,3,8-trihydroxy-6-methyl-anthraquinone) is a moderately potent and poorly selective inhibitor of CK2, one of the most pleiotropic Ser/Thr protein kinases, implicated in neoplasia and in other global diseases. By virtual screening of the MMS database we have now identified quinalizarin (1,2,5,8-tetrahydroxy-anthraquinone) as an inhibitor of CK2 more potent and selective than emodin. CK2 inhibition by quinalizarin is competitive with respect to ATP, with a Ki value around 50 nM. Tested at 1 μM concentration on a panel of 75 protein kinases, quinalizarin drastically inhibits only CK2, with a promiscuity score ...
Source: BJ Structure - May 11, 2009 Category: Biochemistry Authors: G Cozza, M Mazzorana, E Papinutto, J Bain, M Elliott, G Di Maira, A Gianoncelli, M A. Pagano, S Sarno, M Ruzzene, R Battistutta, F Meggio, S Moro, G Zagotto, L A. Pinna Tags: BJ BJ Signal Source Type: journals

NEMO oligomerisation and its ubiquitin-binding propertiesEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
The IKK complex is a key regulatory component of NF-κB activation and is responsible for mediating the degradation of IκB, thereby allowing nuclear translocation of NF-κB and transcription of target genes. NEMO, the regulatory subunit of the IKK complex plays a pivotal role in this process by integrating upstream signals, in particular the recognition of poly-ubiquitin chains, and relaying these to the activation of IKKα and IKKβ, the catalytic subunits of the IKK complex. The oligomeric state of NEMO is controversial and the mechanism by which it regulates activation of the IKK complex i...
Source: BJ Structure - May 7, 2009 Category: Biochemistry Authors: F J Ivins, M G Montgomery, S JM Smith, A C Morris-Davies, I A Taylor, K Rittinger Tags: BJ BJ Structure Source Type: journals

Kinetic analysis of human protein arginine N-methyltransferase 2: Formation of monomethyl- and asymmetric dimethylarginine residues on histone H4Email this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Protein arginine N-methyltransferases (PRMTs) methylate arginine residues within proteins using S-adenosyl-L-methionine (AdoMet) to form S-adenosyl-L-homocysteine and methylarginine residues. All PRMTs produce ω-NG-monomethylarginine (MMA) residues and either asymmetric ω-NG-,NG--dimethylarginine (aDMA) or symmetric ω-NG-,N’G--dimethylarginine (sDMA) residues, referred to as Type I or Type II activity, respectively. Here we report methylation activity from PRMT2 and compare it to PRMT1 activity using ultrahigh-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS), gel electroph...
Source: BJ Structure - April 30, 2009 Category: Biochemistry Authors: T M. Lakowski, A Frankel Tags: BJ BJ Structure Source Type: journals

Pseudomonas aeruginosa MurE amide ligase: enzyme kinetics and peptide inhibitorEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
The enzyme kinetics of the amide ligase MurE from Pseudomonas aeruginosa were determined using the synthesized nucleotide substrate UDP-MurNAc-Ala-Glu. When coupled to a competitive bio-panning using a M13 phage display library encoding ~ 2.7 x 109 random peptide permutations and the specific substrates meso-A2pm and ATP, a peptide inhibitor of MurE was identified. The MurEp1 dodecamer selected and synthesized inhibited MurE ATPase activity with an IC50 value of 500 μM. The inhibition was shown to be time-dependent and reversed by the addition of meso-A2pm or UDPMurNAc- Ala-Glu during the pre-incubation step. Kineti...
Source: BJ Structure - April 28, 2009 Category: Biochemistry Authors: C Paradis-Bleau, A Lloyd, F Sanschagrin, H Maaroufi, T Clarke, A Blewett, C Dowson, D I. Roper, T D. H. Bugg, R C. Levesque Tags: BJ Structure Source Type: journals

Receptor associated protein (RAP) has two high affinity binding sites for the low density lipoprotein receptor-related protein (LRP). Consequences for the chaperone functions of RAPEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Receptor associated protein (RAP) is a three domain 38 kDa ER-resident protein that is a chaperone for the low density lipoprotein receptor-related protein (LRP). Whereas RAP is known to compete for binding of all known LRP ligands, neither the location, the number of binding sites on LRP, nor the domains of RAP involved in binding is known with certainty. We have systematically examined the binding of each of the three RAP domains (D1, D2 and D3) to tandem and triple complement-like repeats (CR) that span the principal ligand binding region, cluster II, of LRP. We found that D3 binds with low nanomolar affinity to all (CR...
Source: BJ Structure - April 27, 2009 Category: Biochemistry Authors: J K. Jensen, K Dolmer, C Schar, P G.W. Gettins Tags: BJ Structure Source Type: journals

Functional characterization of LePGT1, a membrane-bound prenyltransferase involved in the geranylation of para-hydroxybenzoic acidEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
The aromatic substrate prenyltransferase (AS-PT) family plays a critical role in the biosynthesis of important quinone compounds such as ubiquinone and plastoquinone, although biochemical characterizations of AS-PTs have rarely been carried out because most members are membrane-bound enzymes with multiple transmembrane α-helices. Para-hydroxybenzoic acid prenyltransferases (PPTs) are a large subfamily of AS-PTs involved in ubiquinone and naphthoquinone biosynthesis. LePGT1 is the regulatory enzyme for the biosynthesis of shikonin, a naphthoquinone pigment, and was utilized in this study as representative of membrane...
Source: BJ Structure - April 24, 2009 Category: Biochemistry Authors: K Ohara, A Muroya, N Fukushima, K Yazaki Tags: BJ Plant Source Type: journals

The catalytic mechanism of NADH-dependent reduction of 9,10-phenanthrenequinone by Candida tenuis xylose reductase reveals plasticity in an aldo-keto reductase active siteEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Despite their widely varying physiological functions in carbonyl metabolism, Candida tenuis xylose reductase (AKR2B5) and many related enzymes of the aldo-keto reductase protein superfamily utilise 9,10 phenanthrenequinone (PQ) as a common in vitro substrate for NAD(P)H-dependent reduction. The catalytic roles of the conserved active-site residues (Tyr51, Lys80, His113) of AKR2B5 in the conversion of the reactive α-dicarbonyl moiety of PQ are not well understood. Using wild-type and mutated (Tyr51, Lys80 and His113 individually replaced by Ala) forms of AKR2B5, we have conducted steady-state and transient kinetic st...
Source: BJ Structure - April 15, 2009 Category: Biochemistry Authors: S L. Pival, M Klimacek, B Nidetzky Tags: BJ Structure Source Type: journals

Structure-activity analysis of aging and reactivation of human butyrylcholinesterase inhibited by analogues of tabunEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
This study provides structural basis to design new oximes capable of reactivating phosphoramidyl-hBChE conjugates after intoxication, notably when hBChE is used as pretreatment or to design BChE-based catalytic bioscavengers. (Source: BJ Structure)
Source: BJ Structure - April 15, 2009 Category: Biochemistry Authors: E Carletti, N Aurbek, E Gillon, M Loiodice, Y Nicolet, J Fontecilla-Camps, P Masson, H Thiermann, F Nachon, F Worek Tags: BJ Structure Source Type: journals

The presence of mono-glucosylated N196-glycan is important for the structural stability of storage protein, arylphorinEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
In this study, we determined the crystal structure of APA and all sugar moieties of the N196-glycan were clearly observed in the electron-density map. Although the sugar moieties of the glycan generally have high structural flexibility, most sugar moieties of the N196-glycan were well organized in the deep cleft of the subunit interface, and mediated many inter- and intra-subunit H-bonds. Analytical ultracentrifugation and guanidinium chloride (GdmCl) unfolding experiments revealed that the presence of the N196-glycan was important for stabilizing the hexameric state and overall stability of APA, respectively. Our results ...
Source: BJ Structure - April 9, 2009 Category: Biochemistry Authors: K Ryu, J Lee, T Kwon, H Choi, H Park, S Hwang, Z Lee, K Lee, Y Han, Y Choi, Y Jeon, C Cheong, S Kim Tags: BJ Structure Source Type: journals

Thermodynamic characterization of the redox centers in a representative domain of a novel c-type multiheme cytochromeEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Multiheme cytochromes that could form protein “nanowires” were identified in Geobacter sulfurreducens genome and represent a new type of multiheme cytochromes. The sequences of these proteins, two with 12-hemes (GSU1996, GSU0592) and one with 27-hemes (GSU2210) suggest that they are formed by domains homologous to triheme cytochromes c7. While all three hemes have bis-His coordination in cytochromes c7, in each domain of the above polymers the heme equivalent to heme IV has His-Met coordination. We previously determined the structure and measured the macroscopic redox potential of one representative domain (d...
Source: BJ Structure - April 8, 2009 Category: Biochemistry Authors: L Morgado, A P Fernandes, Y Y Londer, P Raj Pokkuluri, M Schiffer, C A Salgueiro Tags: BJ Energy Source Type: journals

Molecular basis of the inhibitor selectivity and insights into the feedback inhibition mechanism of citramalate synthase from Leptospira interrogansEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
Citramalate synthase (LiCMS) catalyzes the first reaction of the isoleucine biosynthesis pathway in Leptospira interrogan, the pathogen of leptospirosis. The catalytic reaction is regulated through feedback inhibition by its end-product isoleucine. To understand the molecular basis of the high selectivity of inhibitor and the mechanism of feedback inhibition, we determined the crystal structure of the regulatory domain of LiCMS (LiCMSC) in complex with Ile and performed biochemical study of the inhibition of LiCMS using mutagenesis and kinetics methods. LiCMSC forms a dimer of dimers in both the crystal structure and solut...
Source: BJ Structure - April 7, 2009 Category: Biochemistry Authors: P Zhang, J Ma, Z Zhang, M Zha, H Xu, G Zhao, J Ding Tags: BJ Structure Source Type: journals

Stability of the two wings of the coiled-coil domain of ClpB chaperone is critical for its disaggregation activityEmail this article to a colleague. Save this article to My Clippings. Discuss or comment on this article.
The ClpB chaperone forms a hexamer ring and rescues aggregated proteins in cooperation with the DnaK system. Each subunit of ClpB has two nucleotide-binding modules, AAA-1 and AAA-2, and an 85-Å-long coiled-coil. The coiled-coil consists of two halves: wing-1, leaning toward AAA-1, and wing-2, leaning away from all the domains. The coiled-coil is stabilized by leucine zipper-like interactions between Leu and Ile residues of two amphipathic α-helices that twist around each other to form each wing. To destabilize the two wings, we developed a series of mutants by replacing these residues with Ala. As the number...
Source: BJ Structure - April 7, 2009 Category: Biochemistry Authors: Y Watanabe, Y Nakazaki, R Suno, M Yoshida Tags: BJ Structure Source Type: journals