Frederic Vellieux, B.Sc., Ph.D., HdR - 1st Faculty of Medicine, Charles University - Dr

Frederic Vellieux

B.Sc., Ph.D., HdR

1st Faculty of Medicine, Charles University

Dr

Vestec, Central Bohemia | Czech Republic

Main Specialties: Chemistry, Pharmacology

Additional Specialties: Structural Biology

ORCID logohttps://orcid.org/0000-0002-1922-5707


Top Author

Frederic Vellieux, B.Sc., Ph.D., HdR - 1st Faculty of Medicine, Charles University - Dr

Frederic Vellieux

B.Sc., Ph.D., HdR

Introduction

09/2018 - current: senior researcher, 1st Faculty of Medicine, Charles University in Prague (BIOCEV Vestec site, CZ)
01/2016 - 12/2018: support man. (scientific manager), Centre of Molecular Structure, Inst. Biotechnology CAS (BIOCEV, Vestec, CZ)
04/1991 - 12/2015: postdoc CEA, Institut de Biologie Structurale CEA CNRS UJF (Grenoble, FR)
10/1988 - 03/1991: postdoc, BIOSON Research Institute, Rijksuniversiteit Groningen (Groningen, NL)
09/1984 - 08/1988: Ph.D. student, Chemical Physics, Rijksuniversiteit Groningen (Groningen, NL)
09/1980 - 06/1984: B.Sc. student (Chemistry & Crystallography), Birkbeck, University of London (London, UK)
10/1977 - 06/1980: Medical student, Faculty of Medicine, University of Angers (Angers, FR)

Primary Affiliation: 1st Faculty of Medicine, Charles University - Vestec, Central Bohemia , Czech Republic

Specialties:

Additional Specialties:

Research Interests:


View Frederic Vellieux’s Resume / CV

Education

Oct 2006
Université de Grenoble
Habilitation
Physics
Jun 1990
Rijksuniversiteit Groningen
Ph.D. (doktoraat)
Protein Crystallography
Jun 1984
Birkbeck, University of London
B. Sc. 1st class hons
Chemistry

Experience

Sep 2018
senior researcher
research
1st Faculty of Medicine, Charles U (until present)
Jan 2016
support man (scientific manager)
research support
Centre of Molecular Structure, IBT (until end Dec 2018)
Apr 1991
postdoc (French Atomic Energy Commission)
research
Institut de Biologie Structurale (until end Dec 2015)
Oct 1988
postdoc (Rijksuniversiteit Groningen)
research
BIOSON Research Institute (until end Mar 1991)
Sep 1984
Ph.D. student (Rijksuniversiteit Groningen)
research
Dept of Chemical Physics (until Sept 1988)

Publications

41Publications

390Reads

200Profile Views

1746PubMed Central Citations

Highly selective mitochondrial probes based on fluorinated pentamethinium salts: On two-photon properties and microscopic applications

Dyes Pig. Jan 2020, 172:107802.

Dyes and Pigments Volume 172, 107802

AbstractFluorescent dyes and probes have been of interest in the fields of chemistry and biology for a long time. However, with more recent applications in advanced techniques, such as two-photon and super-resolution fluorescence microscopy, higher demands are being placed on physico-chemical properties of such compounds. Another requirement is specific intracellular targeting which often remains a big challenge. Here, we present the synthesis of a series of far red-emitting fluorescent dyes based on a structure of fluorinated symmetrical ɣ-aryl substituted pentamethinium salts. We reveal the relationship between the structure and photophysical and biological properties of the compounds; a type of a side unit (indole, benzothiazole) and presence of fluorine atoms were investigated. It was found that substitution of benzothiazolium side units for indolium ones led to increased quantum yields and higher photostability of the dyes and that the presence of fluorine on the γ-aryl moiety did not exhibit significant influence on their photophysical properties. Regardless of the structural difference, all compounds localized in mitochondria of various cancerous and noncancerous cell lines and can be utilized in live-cell imaging using wide-field, confocal, two-photon and structured illumination fluorescence microscopy. Our findings suggest that the properties of symmetrical pentamethinium salts are affected to a higher extent by the type of a side unit rather than fluorine atoms on the γ-aryl moiety. Last but not least, we believe that such probes will further extend tools for mitochondrial research.

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January 2020

Impact Factor 3.767

26 Reads

Synthesis, Conformational Analysis and Crystal Structure of New Thioxo, Oxo, Seleno Diastereomeric Cyclophosphamides Containing 1,3,2-dioxaphosphorinane

Current Organic Chemistry 2019, DOI : 10.2174/1385272823666190213142748

Current Organic Chemistry

Abstract: Thioxo, Oxo, Seleno and diastereomeric cyclophosphamides containing 1,3,2-dioxaphosphorinane are prepared by a one-step chemical reaction. Their structural determination is carried out by means of Nuclear Magnetic Resonance NMR (31P, 1 H, 13C) and High-Resolution Mass Spectroscopy (HRMS). The conformational study of diastereomeric products is described. Density Functional Theory (DFT) calculations allowed the identification of preferred conformations. Experimental and calculated 31P, 13C, 1H NMR chemical shifts are compared. The molecular structure of the 2-Benzylamino-5-methyl-5-propyl-2-oxo-1,3,2-dioxaphosphorinane (3d) has been determined by means of crystal X-ray diffraction methods.

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February 2019

Impact Factor 2.193

122 Reads

Crystal structure of pb9, the distal tail protein of bacteriophage T5: a conserved structural motif among all siphophages.

J Virol 2014 Jan 23;88(2):820-8. Epub 2013 Oct 23.

Univ. Grenoble Alpes, Institut de Biologie Structurale (IBS), Grenoble, France.

The tail of Caudovirales bacteriophages serves as an adsorption device, a host cell wall-perforating machine, and a genome delivery pathway. In Siphoviridae, the assembly of the long and flexible tail is a highly cooperative and regulated process that is initiated from the proteins forming the distal tail tip complex. In Gram-positive-bacterium-infecting siphophages, the distal tail (Dit) protein has been structurally characterized and is proposed to represent a baseplate hub docking structure. It is organized as a hexameric ring that connects the tail tube and the adsorption device. In this study, we report the characterization of pb9, a tail tip protein of Escherichia coli bacteriophage T5. By immunolocalization, we show that pb9 is located in the upper part of the cone of the T5 tail tip, at the end of the tail tube. The crystal structure of pb9 reveals a two-domain protein. Domain A exhibits remarkable structural similarity with the N-terminal domain of known Dit proteins, while domain B adopts an oligosaccharide/oligonucleotide-binding fold (OB-fold) that is not shared by these proteins. We thus propose that pb9 is the Dit protein of T5, making it the first Dit protein described for a Gram-negative-bacterium-infecting siphophage. Multiple sequence alignments suggest that pb9 is a paradigm for a large family of Dit proteins of siphophages infecting mostly Gram-negative hosts. The modular structure of the Dit protein maintains the basic building block that would be conserved among all siphophages, combining it with a more divergent domain that might serve specific host adhesion properties.

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http://dx.doi.org/10.1128/JVI.02135-13DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3911636PMC
January 2014
12 Reads
30 Citations
4.439 Impact Factor

Using lanthanoid complexes to phase large macromolecular assemblies.

J Synchrotron Radiat 2011 Jan 5;18(1):74-8. Epub 2010 Nov 5.

CEA, IBS, F-38054 Grenoble, France.

Lanthanoid ions exhibit extremely large anomalous X-ray scattering at their L(III) absorption edge. They are thus well suited for anomalous diffraction experiments. A novel class of lanthanoid complexes has been developed that combines the physical properties of lanthanoid atoms with functional chemical groups that allow non-covalent binding to proteins. Two structures of large multimeric proteins have already been determined by using such complexes. Here the use of the luminescent europium tris-dipicolinate complex [Eu(DPA)(3)](3-) to solve the low-resolution structure of a 444 kDa homododecameric aminopeptidase, called PhTET1-12s from the archaea Pyrococcus horikoshii, is reported. Surprisingly, considering the low resolution of the data, the experimental electron density map is very well defined. Experimental phases obtained by using the lanthanoid complex lead to maps displaying particular structural features usually observed in higher-resolution maps. Such complexes open a new way for solving the structure of large molecular assemblies, even with low-resolution data.

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http://dx.doi.org/10.1107/S0909049510036824DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3004260PMC
January 2011
7 Reads
11 Citations
2.736 Impact Factor

The structural and biochemical characterizations of a novel TET peptidase complex from Pyrococcus horikoshii reveal an integrated peptide degradation system in hyperthermophilic Archaea.

Mol Microbiol 2009 Apr 5;72(1):26-40. Epub 2009 Mar 5.

Institut de Biologie Structurale J.-P. Ebel, UMR 5075 CNRS-CEA-UJF, 41 rue Jules Horowitz, 38027 Grenoble, France.

The structure of a 468 kDa peptidase complex from the hyperthermophile Pyrococcus horikoshii has been solved at 1.9 A resolution. The monomer contains the M42 peptidase typical catalytic domain, and a dimerization domain that allows the formation of dimers that assemble as a 12-subunit self-compartmentalized tetrahedron, similar to those described for the TET peptidases. The biochemical analysis shows that the enzyme is cobalt-activated and cleaves peptides by a non-processive mechanism. Consequently, this protein represents the third TET peptidase complex described in P. horikoshii, thereby called PhTET3. It is a lysyl aminopeptidase with a strong preference for basic residues, which are poorly cleaved by PhTET1 and PhTET2. The structural analysis of PhTET3 and its comparison with PhTET1 and PhTET2 unravels common features explaining the general mode of action of the TET molecular machines as well as differences that can be associated with strong substrate discriminations. The question of the stability of the TET assemblies under extreme temperatures has been addressed. PhTET3 displays its maximal activity at 95 degrees C and small-angle neutron scattering experiments at 90 degrees C demonstrate the absence of quaternary structure alterations after extensive incubation times. In conclusion, PhTETs are complementary peptide destruction machines that may play an important role in the metabolism of P. horikoshii.

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http://doi.wiley.com/10.1111/j.1365-2958.2009.06600.x
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http://dx.doi.org/10.1111/j.1365-2958.2009.06600.xDOI Listing
April 2009
9 Reads
32 Citations
4.419 Impact Factor

Noncrystallographic symmetry averaging in phase refinement and extension.

Methods Enzymol 1997 ;277:18-53

Institut de Biologie Structurale, Jean-Pierre Ebel CEA CNRS, Grenoble 01, France.

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http://dx.doi.org/10.1016/s0076-6879(97)77005-8DOI Listing
May 2008
77 Reads
46 Citations
2.088 Impact Factor

Activity, stability and structural studies of lactate dehydrogenases adapted to extreme thermal environments.

J Mol Biol 2007 Nov 22;374(2):547-62. Epub 2007 Sep 22.

Laboratoire de Biophysique Moléculaire, Institut de Biologie Structurale J.-P. Ebel, CEA CNRS UJF, UMR 5075, 41 rue Jules Horowitz, 38027 Grenoble Cedex 01, France.

Lactate dehydrogenase (LDH) catalyzes the conversion of pyruvate to lactate with concomitant oxidation of NADH during the last step in anaerobic glycolysis. In the present study, we present a comparative biochemical and structural analysis of various LDHs adapted to function over a large temperature range. The enzymes were from Champsocephalus gunnari (an Antarctic fish), Deinococcus radiodurans (a mesophilic bacterium) and Thermus thermophilus (a hyperthermophilic bacterium). The thermodynamic activation parameters of these LDHs indicated that temperature adaptation from hot to cold conditions was due to a decrease in the activation enthalpy and an increase in activation entropy. The crystal structures of these LDHs have been solved. Pairwise comparisons at the structural level, between hyperthermophilic versus mesophilic LDHs and mesophilic versus psychrophilic LDHs, have revealed that temperature adaptation is due to a few amino acid substitutions that are localized in critical regions of the enzyme. These substitutions, each having accumulating effects, play a role in either the conformational stability or the local flexibility or in both. Going from hot- to cold-adapted LDHs, the various substitutions have decreased the number of ion pairs, reduced the size of ionic networks, created unfavorable interactions involving charged residues and induced strong local disorder. The analysis of the LDHs adapted to extreme temperatures shed light on how evolutionary processes shift the subtle balance between overall stability and flexibility of an enzyme.

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http://dx.doi.org/10.1016/j.jmb.2007.09.049DOI Listing
November 2007
3 Reads
112 Citations
4.333 Impact Factor

Molecular Adaptation to High Salt

Phys. Biochem. Extremophiles 2007:240

Physiology and Biochemistry of Extremophiles

IntroductionHalophilic organisms inhabit extremely saline environments up to NaCl saturation. Halophiles are found in all three domains of life: Bacteria, Archaea, and Eukarya. One of the motivations for their study is the hope to reach an understanding of the molecular and cellular mechanisms underlying their ability to cope with these hostile conditions. Another motivation comes from the fact that a large number of halophilic microorganisms are Archaea, and some macromolecular machineries from Archaea share similarities with those from Eukarya. Examples are complexes involved in translation, proteolysis, or protein folding (Langer et al., 1995; Maupin-Furlow et al., 2004). Archaea, therefore, offer simple macromolecular models to describe systems that are more complex in Eukarya.Extreme halophiles require multimolar salt for growth. Their study has shown that they have developed a wide variety of strategies to thrive in media that are hostile to other life forms (for a complete review, see Oren, 2002). In order to counterbalance the external osmotic pressure, extreme halophiles accumulate salt—mainly KCl—close to saturation in their cytosol. All biochemical reactions occur in this extreme medium.In this chapter, we first present briefly the insights into adaptation provided by the study of the four genomes of extreme halophiles sequenced to date. The focus will then shift to molecular adaptation of halophilic proteins, defined as proteins isolated from extreme halophiles. We shall not address membrane proteins or ribosomes. The starting point of the analysis will be the high-resolution structures of halophilic proteins available at this time. DNA–protein interactions will be considered with the only example described so far, which concerns DNA binding a protein from a nonextreme halophile. Structural information has been combined with complementary phylogenetic analysis and solution studies. Different aspects concerning solvation, stabilization of the folded and associated assemblies of proteins, and salt effect will be presented. Molecular evolution also has to select appropriate solubility and dynamics in order to permit and favor halophilic protein activity at high salt.

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August 2007

5 Citations

Specific radiation damage to acidic residues and its relation to their chemical and structural environment.

J Synchrotron Radiat 2007 Jan 15;14(Pt 1):84-91. Epub 2006 Dec 15.

Laboratoire de Biophysique Moléculaire, Institut de Biologie Structurale J.-P. Ebel, CEA CNRS UJF, 41 Rue Jules Horowitz, 38027 Grenoble CEDEX 1, France.

Intense synchrotron radiation produces specific structural and chemical damage to crystalline proteins even at 100 K. Carboxyl groups of acidic residues (Glu, Asp) losing their definition is one of the major effects observed. Here, the susceptibilities to X-ray damage of acidic residues in tetrameric malate dehydrogenase from Haloarcula marismortui are investigated. The marked excess of acidic residues in this halophilic enzyme makes it an ideal target to determine how specific damage to acidic residues is related to their structural and chemical environment. Four conclusions are drawn. (i) Acidic residues interacting with the side-chains of lysine and arginine residues are less affected by radiation damage than those interacting with serine, threonine and tyrosine side-chains. This suggests that residues with higher pK(a) values are more vulnerable to damage than those with a lower pK(a). However, such a correlation was not found when calculated pK(a) values were inspected. (ii) Acidic side-chains located in the enzymatic active site are the most radiation-sensitive ones. (iii) Acidic residues in the internal cavity formed by the four monomers and those involved in crystal contacts appear to be particularly susceptible. (iv) No correlation was found between radiation susceptibility and solvent accessibility.

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http://dx.doi.org/10.1107/S0909049506038623DOI Listing
January 2007
25 Reads
66 Citations
2.736 Impact Factor

An archaeal peptidase assembles into two different quaternary structures: A tetrahedron and a giant octahedron.

J Biol Chem 2006 Nov 14;281(47):36327-37. Epub 2006 Sep 14.

Laboratoire de Virologie Moléculaire et Structurale c/o EMBL, FRE 2854 CNRS-UJF, 38042 Grenoble, France.

Cellular proteolysis involves large oligomeric peptidases that play key roles in the regulation of many cellular processes. The cobalt-activated peptidase TET1 from the hyperthermophilic Archaea Pyrococcus horikoshii (PhTET1) was found to assemble as a 12-subunit tetrahedron and as a 24-subunit octahedral particle. Both quaternary structures were solved by combining x-ray crystallography and cryoelectron microscopy data. The internal organization of the PhTET1 particles reveals highly self-compartmentalized systems made of networks of access channels extended by vast catalytic chambers. The two edifices display aminopeptidase activity, and their organizations indicate substrate navigation mechanisms different from those described in other large peptidase complexes. Compared with the tetrahedron, the octahedron forms a more expanded hollow structure, representing a new type of giant peptidase complex. PhTET1 assembles into two different quaternary structures because of quasi-equivalent contacts that previously have only been identified in viral capsids.

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http://dx.doi.org/10.1074/jbc.M604417200DOI Listing
November 2006
15 Reads
38 Citations
4.573 Impact Factor

Methanoarchaeal sulfolactate dehydrogenase: prototype of a new family of NADH-dependent enzymes.

EMBO J 2004 Mar 11;23(6):1234-44. Epub 2004 Mar 11.

Laboratoire de Biophysique Moléculaire, Institut de Biologie Structurale J-P Ebel CEA CNRS UJF, Grenoble, France.

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http://emboj.embopress.org/cgi/doi/10.1038/sj.emboj.7600147
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http://dx.doi.org/10.1038/sj.emboj.7600147DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC381418PMC
March 2004
6 Reads
19 Citations
10.434 Impact Factor

The 2.9A resolution crystal structure of malate dehydrogenase from Archaeoglobus fulgidus: mechanisms of oligomerisation and thermal stabilisation.

J Mol Biol 2004 Jan;335(1):343-56

Laboratoire de Biophysique Moléculaire, Institut de Biologie Structurale J.-P. Ebel CEA CNRS UJF, UMR-5075, 41 rue Jules Horowitz, 38027 Cedex 01, Grenoble, France.

The crystal structure of malate dehydrogenase from the hyperthermophilic archaeon Archeoglobus fulgidus, in complex with its cofactor NAD, was solved at 2.9A resolution. The crystal structure shows a compact homodimer with one coenzyme bound per subunit. The substrate binding site is occupied by a sulphate ion. In order to gain insight into adaptation mechanisms, which allow the protein to be stable and active at high temperatures, the 3D structure was compared to those of several thermostable and hyperthermostable homologues, and to halophilic malate dehydrogenase. The hyperthermostable A. fulgidus MalDH protein displays a reduction of the solvent-exposed surface, an optimised compact hydrophobic core, a high number of hydrogen bonds, and includes a large number of ion pairs at the protein surface. These features occur concomitantly with a reduced number of residues in the protein subunit, due to several deletions in loop regions. The loops are further stiffened by ion pair links with secondary structure elements. A. fulgidus malate dehydrogenase is the only dimeric protein known to date that belongs to the [LDH-like] MalDH family. All the other known members of this family are homo-tetramers. The crystal structures revealed that the association of the dimers to form tetramers is prevented by several deletions, taking place at the level of two loops that are known to be essential for the tetramerisation process within the LDH and [LDH-like] MalDH enzymes.

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http://dx.doi.org/10.1016/j.jmb.2003.10.054DOI Listing
January 2004
7 Reads
31 Citations
4.333 Impact Factor

The Oligomeric states of Haloarcula marismortui malate dehydrogenase are modulated by solvent components as shown by crystallographic and biochemical studies.

J Mol Biol 2003 Feb;326(3):859-73

Laboratoire de Biophysique Moléculaire, Institut de Biologie Structurale J.-P. Ebel CEA CNRS UJF UMR-5075, 41 rue Jules Horowitz, 38027 Grenoble Cedex 01, France.

The three-dimensional crystal structure of the (R207S, R292S) mutant of malate dehydrogenase from Haloarcula marismortui was solved at 1.95A resolution in order to determine the role of salt bridges and solvent ions in halophilic adaptation and quaternary structure stability. The mutations, located at the dimer-dimer interface, disrupt two inter-dimeric salt bridge clusters that are essential for wild-type tetramer stabilisation. Previous experiments in solution, performed on the double mutant, had shown a tetrameric structure in 4M NaCl, which dissociated into active dimers in 2M NaCl. In order to establish if the active dimeric form is a product of the mutation, or if it also exists in the wild-type protein, complementary studies were performed on the wild-type enzyme by analytical centrifugation and small angle neutron scattering experiments. They showed the existence of active dimers in NaF, KF, Na(2)SO(4), even in the absence of NADH, and in the presence of NADH at concentrations of NaCl below 0.3M. The crystal structure shows a tetramer that, in the absence of the salt bridge clusters, appears to be stabilized by a network of ordered water molecules and by Cl(-) binding at the dimer-dimer interface. The double mutant and wild-type dimer folds are essentially identical (the r.m.s. deviation between equivalent C(alpha) positions is 0.39A). Chloride ions are also observed at the monomer-monomer interfaces of the mutant, contributing to the stability of each dimer against low salt dissociation. Our results support the hypothesis that extensive binding of water and salt is an important feature of adaptation to a halophilic environment.

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http://dx.doi.org/10.1016/s0022-2836(02)01450-xDOI Listing
February 2003
4 Reads
59 Citations
4.333 Impact Factor

The 3.0 A resolution crystal structure of glycosomal pyruvate phosphate dikinase from Trypanosoma brucei.

J Mol Biol 2002 May;318(5):1417-32

Laboratoire de Biophysique Moleculaire, Institut de Biologie Structurale J.-P. Ebel CEA CNRS UJF, Grenoble, France.

The crystal structure of the glycosomal enzyme pyruvate phosphate dikinase from the African protozoan parasite Trypanosoma brucei has been solved to 3.0 A resolution by molecular replacement. The search model was the 2.3 A resolution structure of the Clostridium symbiosum enzyme. Due to different relative orientations of the domains and sub-domains in the two structures, molecular replacement could be achieved only by positioning these elements (four bodies altogether) sequentially in the asymmetric unit of the P2(1)2(1)2 crystal, which contains one pyruvate phosphate dikinase (PPDK) subunit. The refined model, comprising 898 residues and 188 solvent molecules per subunit, has a crystallographic residual index Rf = 0.245 (cross-validation residual index Rfree = 0.291) and displays satisfactory stereochemistry. Eight regions, comprising a total of 69 amino acid residues at the surface of the molecule, are disordered in this crystal form. The PPDK subunits are arranged around the crystallographic 2-fold axis as a dimer, analogous to that observed in the C. symbiosum enzyme. Comparison of the two structures was carried out by superposition of the models. Although the fold of each domain or sub-domain is similar, the relative orientations of these constitutive elements are different in the two structures. The trypanosome enzyme is more "bent" than the bacterial enzyme, with bending increasing from the center of the molecule (close to the molecular 2-fold axis) towards the periphery where the N-terminal domain is located. As a consequence of this increased bending and of the differences in relative positions of subdomains, the nucleotide-binding cleft in the amino-terminal domain is wider in T. brucei PPDK: the N-terminal fragment of the amino-terminal domain is distant from the catalytic, phospho-transfer competent histidine 482 (ca 10 A away). Our observations suggest that the requirements of domain motion during enzyme catalysis might include widening of the nucleotide-binding cleft to allow access and departure of the AMP or ATP ligand.

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http://dx.doi.org/10.1016/s0022-2836(02)00113-4DOI Listing
May 2002
13 Reads
26 Citations
4.333 Impact Factor

Crystallization and preliminary crystallographic investigation of glycosomal pyruvate phosphate dikinase from Trypanosoma brucei.

Acta Crystallogr D Biol Crystallogr 2000 Dec;56(Pt 12):1688-90

Laboratoire de Biophysique Moléculaire, Institut de Biologie Structurale J.-P. Ebel CEA CNRS, 41 Rue Jules Horowitz, 38027 Grenoble CEDEX 01, France.

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http://dx.doi.org/10.1107/s0907444900015298DOI Listing
December 2000
4 Reads
7.232 Impact Factor

Crystallographic studies of the interaction between the ferredoxin-NADP+ reductase and ferredoxin from the cyanobacterium Anabaena: looking for the elusive ferredoxin molecule.

Acta Crystallogr D Biol Crystallogr 2000 Nov;56(Pt 11):1408-12

LCCP, Institut de Biologie Structurale J. P. Ebel, CEA-CNRS, 41 Rue Jules Horowitz, F38027 Grenoble, France.

Ferredoxin-NADP(+) reductase (FNR) and its physiological electron donor ferredoxin (Fd) from the cyanobacterium Anabaena PCC7119 have been co-crystallized. The unit-cell parameters are a = b = 63.72, c = 158.02 A and the space group is P2(1)2(1)2(1). The crystal structure has been solved with 2.4 A resolution synchrotron data by molecular replacement, anomalous dispersion and R(min) search methods. For the computations, the crystal was treated as a merohedral twin. The asymmetric unit contains two FNR molecules and one ferredoxin molecule. The packing of the FNR molecules displays a nearly tetragonal symmetry (space group P4(3)2(1)2), whereas the ferredoxin arrangement is orthorhombic. This study provides the first crystallographic model of a dissociable complex between FNR and Fd.

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http://dx.doi.org/10.1107/s0907444900010052DOI Listing
November 2000
3 Reads
44 Citations
7.232 Impact Factor

PBR: a heavy-atom refinement and phasing procedure to reduce phase bias when heavy-atom derivatives contain common sites.

Acta Crystallogr D Biol Crystallogr 1999 Feb;55(Pt 2):469-72

Laboratoire de Cristallographie et de Cristallogenèse des Protéines, Institut de Biologie Structurale J.-P. Ebel CEA--CNRS, 41 Avenue des Martyrs, 38027 Grenoble CEDEX 01, France.

A procedure, called PBR (phase-bias reduction), has been developed to properly refine heavy-atom derivatives and to generate less biased heavy-atom phases when these derivatives contain common heavy-atom sites. Two independent events are obtained by splitting the refinement and phasing calculations into two stages, the first in which one of the derivatives having common sites is used together with the native amplitudes and the second in which both derivatives with common sites are used simultaneously, with one of them being used as the native data set. Improved centroid phases and the corresponding figures of merit are obtained by phase combination. This procedure has been used in the structure determination of the iron-cluster-containing protein -pyruvate-ferredoxin oxidoreductase. When the common heavy-atom sites are properly treated by the PBR procedure, the resulting calculated centroid phases are improved with respect to classical heavy-atom refinement centroid phases where all derivatives are refined together. This leads to improved electron-density distributions, since anomalous difference Fourier maps calculated with the PBR-refined centroid phases and corresponding figures of merit show more clearly the positions of the iron sites.

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http://dx.doi.org/10.1107/s0907444998011366DOI Listing
February 1999
3 Reads
3 Citations
7.232 Impact Factor

A comparison of two algorithms for electron-density map improvement by introduction of atomicity: skeletonization, and map sorting followed by refinement.

Authors:
F M Vellieux

Acta Crystallogr D Biol Crystallogr 1998 Jan;54(Pt 1):81-5

Institut de Biologie Structurale J.-P. Ebel CEA CNRS, 41 Avenue des Martyrs, 38027 Grenoble CEDEX 01, France.

A comparison has been made of two methods for electron-density map improvement by the introduction of atomicity, namely the iterative skeletonization procedure of the CCP4 program DM [Cowtan & Main (1993). Acta Cryst. D49, 148-157] and the pseudo-atom introduction followed by the refinement protocol in the program suite DEMON/ANGEL [Vellieux, Hunt, Roy & Read (1995). J. Appl. Cryst. 28, 347-351]. Tests carried out using the 3.0 A resolution electron density resulting from iterative 12-fold non-crystallographic symmetry averaging and solvent flattening for the Pseudomonas aeruginosa ornithine transcarbamoylase [Villeret, Tricot, Stalon & Dideberg (1995). Proc. Natl Acad. Sci. USA, 92, 10762-10766] indicate that pseudo-atom introduction followed by refinement performs much better than iterative skeletonization: with the former method, a phase improvement of 15.3 degrees is obtained with respect to the initial density modification phases. With iterative skeletonization a phase degradation of 0.4 degrees is obtained. Consequently, the electron-density maps obtained using pseudo-atom phases or pseudo-atom phases combined with density-modification phases are much easier to interpret. These tests also show that for ornithine transcarbamoylase, where 12-fold non-crystallographic symmetry is present in the P1 crystals, G-function coupling leads to the simultaneous decrease of the conventional R factor and of the free R factor, a phenomenon which is not observed when non-crystallographic symmetry is absent from the crystal. The method is far less effective in such a case, and the results obtained suggest that the map sorting followed by refinement stage should be by-passed to obtain interpretable electron-density distributions.

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http://dx.doi.org/10.1107/s0907444997008081DOI Listing
January 1998
3 Reads
13 Citations
7.232 Impact Factor

A connected set algorithm for the identification of spatially contiguous regions in crystallographic envelopes.

Acta Crystallogr D Biol Crystallogr 1997 Jul;53(Pt 4):434-7

Howard Hughes Medical Institute and Department of Biochemistry, University of Texas Southwestern Medical Center, Dallas, TX 75235-9050, USA.

A simple algorithm is described for the identification of spatially contiguous regions in crystallographic envelopes. In a single pass through the grid points of the envelope map, the occupied points are assigned to a series of locally contiguous sets based on consideration of the connections within single voxels. A spatially contiguous region is identified as the union of all of the locally contiguous sets that share an element in common. Therefore, chains of spatial connectivity are traced implicitly by performing simple set operations. This algorithm has been implemented in the program CNCTDENV as part of the DEMON/ANGEL suite of density-modification programs.

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http://dx.doi.org/10.1107/S0907444997001431DOI Listing
July 1997
4 Reads
3 Citations
7.232 Impact Factor

A systematic analysis of weighting schemes for structure factors in macromolecular crystallography

Authors:
FM Vellieux

J Appl. Cryst. 1997 Jun 1;30(3):400

Journal of Applied Crystallography

AbstractThe three weighting schemes for structure-factor amplitudes used in macromolecular crystallography, and the products of these weighting schemes, have been analysed by comparison of the figures of merit to the cosine of the phase difference between the current phase estimate and the true phase, and by computation of the linear correlation coefficient between these figures. The results of the analysis indicate that (i) the Sigma-A and SIM weighting schemes are superior to all others investigated (for the latter scheme, provided that SigmaL has been computed by the method of Henderson & Moffat [Acta Cryst. (1971) B27, 1414-1420]), (ii) all weighting schemes are an underestimate of the cosine of the phase difference, (iii) the correlation between the Rayment figures of merit and the cosine of the phase difference is quite low, indicating a poor weighting scheme, and (iv) for the figures of merit obtained by multiplication of weights, better results are obtained when the best weighting schemes, namely the SigmaA and Sim figures of merit, are combined.

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June 1997

2 Citations

Impact Factor 3.422

Computation of Bhat's OMIT maps with different coefficients

J Appl Cryst 1997 Jun 1; 30(3):396

Journal of Applied Crystallography

AbstractThe OMIT electron-density-map calculation of Bhat [Bhat & Cohen (1984)..I. Appl. Cryst. 17, 244-248; Bhat (1988). J. Appl. Cryst. 21,279-281] is very effective in discovering errors in a macromolecular structure determination. A Fortran program called OMIT has been written to calculate such maps and an investigation has been carried out into which coefficients for the map calculation produce the best OMIT maps. Testing of the program on Savinase ~ showed that the best overall results were obtained when lFol without figure of merit was used. In regions where the map is incorrect, the most interesting OMIT maps are produced when only the figure of merit, or modified SIGMAA coefficients, are used as the initial map amplitude coefficients. Thus, these tests suggest that such OMIT maps are particularly useful to reconstruct the macromolecular model in the grossly incorrect regions of the model.

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June 1997

55 Citations

Impact Factor 3.422

X-ray structure of the ferredoxin:NADP+ reductase from the cyanobacterium Anabaena PCC 7119 at 1.8 A resolution, and crystallographic studies of NADP+ binding at 2.25 A resolution.

J Mol Biol 1996 Oct;263(1):20-39

IBS/LCCP, Grenoble, France.

The crystal structure of the ferredoxin:NADP+ reductase (FNR) from the cyanobacterium Anabaena PCC 7119 has been determined at 2.6 A resolution by multiple isomorphous replacement and refined using 15.0 A to 1.8 A data, collected at 4 degrees C, to an R-factor of 0.172. The model includes 303 residues, the flavin adenine dinucleotide cofactor (FAD), one sulfate ion located at the putative NADP+ binding site and 328 water molecule sites. The structure of Anabaena FNR, including FAD, a network of intrinsic water molecules and a large hydrophobic cavity in the C-terminal domain, resembles that of the spinach enzyme. The major differences concern the additional short alpha-helix (residues 172 to 177 in Anabaena FNR) and residues Arg 100 and Arg 233 which binds NADP+ instead of Lys 116 and Lys 244 in the spinach enzyme. Crystals of a complex of Anabaena FNR with NADP+ were obtained. The model of the complex has been refined using 15 A to 2.25 A X-ray data, collected at -170 degrees C, to an R-factor of 0.186. This model includes 295 residues, FAD, the full NADP+ (with an occupancy of 0.8) and 444 water molecules. The 2'-5' adenine moiety of NADP+ binds to the protein as 2'-phospho-5'-AMP to the spinach FNR. The nicotinamide moiety is turned towards the surface of the protein instead of stacking onto the FAD isoalloxazine ring as would be required for hydride transfer. The model of the complex agrees with previous biochemical studies as residues Arg 100 and Arg 233 are involved in NADP+ binding and residues Arg77, Lys 53 and Lys 294, located on the FAD side of the enzyme, remain free to interact with ferredoxin and flavodoxin, the physiological partners of ferredoxin: NADP reductase.

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http://dx.doi.org/10.1006/jmbi.1996.0553DOI Listing
October 1996
4 Reads
158 Citations
4.333 Impact Factor

Crystal structures of a ferredoxin: NADP+ reductase and of a complex with NADP+.

Biochem Soc Trans 1996 Feb;24(1):10S

IBS/LCCP 41, Grenoble, France.

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http://dx.doi.org/10.1042/bst024010sDOI Listing
February 1996
3 Reads
6 Citations
3.194 Impact Factor

Crystal structure of recombinant triosephosphate isomerase from Bacillus stearothermophilus. An analysis of potential thermostability factors in six isomerases with known three-dimensional structures points to the importance of hydrophobic interactions.

Protein Sci 1995 Dec;4(12):2594-604

Department of Biological Structure, School of Medicine, University of Washington, Seattle 98195, USA.

The structure of the thermostable triosephosphate isomerase (TIM) from Bacillus stearothermophilus complexed with the competitive inhibitor 2-phosphoglycolate was determined by X-ray crystallography to a resolution of 2.8 A. The structure was solved by molecular replacement using XPLOR. Twofold averaging and solvent flattening was applied to improve the quality of the map. Active sites in both the subunits are occupied by the inhibitor and the flexible loop adopts the "closed" conformation in either subunit. The crystallographic R-factor is 17.6% with good geometry. The two subunits have an RMS deviation of 0.29 A for 248 C alpha atoms and have average temperature factors of 18.9 and 15.9 A2, respectively. In both subunits, the active site Lys 10 adopts an unusual phi, psi combination. A comparison between the six known thermophilic and mesophilic TIM structures was conducted in order to understand the higher stability of B. stearothermophilus TIM. Although the ratio Arg/(Arg+Lys) is higher in B. stearothermophilus TIM, the structure comparisons do not directly correlate this higher ratio to the better stability of the B. stearothermophilus enzyme. A higher number of prolines contributes to the higher stability of B. stearothermophilus TIM. Analysis of the known TIM sequences points out that the replacement of a structurally crucial asparagine by a histidine at the interface of monomers, thus avoiding the risk of deamidation and thereby introducing a negative charge at the interface, may be one of the factors for adaptability at higher temperatures in the TIM family. Analysis of buried cavities and the areas lining these cavities also contributes to the greater thermal stability of the B. stearothermophilus enzyme. However, the most outstanding result of the structure comparisons appears to point to the hydrophobic stabilization of dimer formation by burying the largest amount of hydrophobic surface area in B. stearothermophilus TIM compared to all five other known TIM structures.

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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2143043PMC
http://dx.doi.org/10.1002/pro.5560041217DOI Listing
December 1995
8 Reads
138 Citations
2.854 Impact Factor

An enveloped cross rotation function

Authors:
FM Vellieux

J Appl. Cryst. 1995 Dec 1; 28(6):834

Journal of Applied Crystallography

AbstractA procedure has been developed for the computation of the cross rotation function in the initial step of molecular replacement. This procedure involves the truncation of the model Patterson map by means of an envelope which follows the shape of this Patterson. Test calculations using bovine phospholipase A2 as the search model and a mutant porcine phospholipase A2 as the target crystal with this enveloped rotation function indicate that the resulting orientation is nearly as that found after Patterson correlation refinement of the solution of the cross rotation function.

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December 1995

3 Citations

Impact Factor 3.422

Refined 3.2 A structure of glycosomal holo glyceraldehyde phosphate dehydrogenase from Trypanosoma brucei brucei.

Acta Crystallogr D Biol Crystallogr 1995 Jul;51(Pt 4):575-89

BIOSON Research Institute, University of Groningen, The Netherlands.

The three-dimensional crystal structure of the enzyme glyceraldehyde phosphate dehydrogenase from the kinetoplastid Trypanosoma brucei brucei has been determined at 3.2 A resolution from a 37% complete data set collected using the Laue method. The crystals used in the structure determination contain one and a half tetrameric enzyme molecules in the asymmetric unit, i.e. six identical subunits. Initial phasing was carried out by the method of molecular replacement using the refined coordinates of holo glyceraldehyde phosphate dehydrogenase from Bacillus stearothermophilus as a search model. The initial electron-density distribution, obtained from the molecular-replacement solution, was greatly improved by a procedure consisting of 36 cycles of iterative non-crystallographic density averaging. During the averaging procedure, the missing reflections (63% of the data) were gradually introduced as map-inversion structure factors. At completion of the procedure, the R-factor between averaged map-inversion amplitudes and observed structure-factor amplitudes was 19.0% for all data between 7.0 and 3.2 A resolution, and that between the map-inversion amplitudes and later recorded structure-factor amplitudes was 41.9%. After model building into the resulting averaged electron-density map, refinement by molecular-dynamics procedures with X-PLOR provided the current model, which has an R-factor of 17.6% for 34 835 reflections between 7.0 and 3.2 A resolution. The refined model, comprising 2735 protein atoms plus one NAD(+) molecule and two sulfate ions per subunit, has r.m.s. deviations from ideality of 0.02 A for bond lengths and 3.6 degrees for bond angles. All subunits, located either within the tetrameric molecule or within the half tetramer present in the asymmetric unit, are related to each other by almost exact twofold symmetry. The overall structure of the glycosomal glyceraldehyde phosphate dehydrogenase subunit and its quaternary arrangement in the tetrameric molecule are similar to that of the enzyme of lobster and Bacillus stearothermophilus (with r.m.s. differences between equivalent Calpha positions of 0.71 and 0.64 A, respectively). The main differences between the structures is the presence of three insertions, plus the substitution of a beta-strand by a short alpha-helix, both occurring at the surface of the glycosomal enzyme subunit.

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http://dx.doi.org/10.1107/S0907444995003015DOI Listing
July 1995
3 Reads
21 Citations
7.232 Impact Factor

DEMON/ANGEL: a suite of programs to carry out density modification

J Appl. Cryst. 1995 Jun 1;28(3):347

Journal of Applied Crystallography

AbstractThe DEMON/ANGEL suite of computer programs has been developed to carry out density modification by non-crystallographic symmetry-averaging, solvent-flattening and histogram-mapping techniques. This suite consists of programs that allow molecular envelopes to be defined and modified, non-crystallographic symmetry operators to be refined either within one crystal form or between several crystal forms, and iterative density modification to be carried out, including non-crystallographic symmetry averaging of electron densities within or between crystal forms.

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June 1995

84 Citations

Impact Factor 3.422

Structural study of ferredoxin-NADP+ reductase from Anabaena PCC 7119 and its complex with NADP+

Flavins Flavoproteins 1993 1994:431

Flavins and Flavoproteins 1993

IntroductionFerredoxin-NADP+ reductase (EC 1.18.1.12, FNR) is a flavoenzyme that plays a key role in the metabolism of photosynthetic organisms by catalyzing the photoreduction of NADP+, using ferredoxin or flavodoxin as electron donors.

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August 1994

3 Citations

Structure of glycosomal glyceraldehyde-3-phosphate dehydrogenase from Trypanosoma brucei determined from Laue data.

Proc Natl Acad Sci U S A 1993 Mar;90(6):2355-9

Department of Chemistry, BIOSON Research Institute, University of Groningen, The Netherlands.

The three-dimensional structure of glycosomal glyceraldehyde-3-phosphate dehydrogenase [D-glyceraldehyde-3-phosphate:NAD+ oxidoreductase (phosphorylating), EC 1.12.1.12] from the sleeping-sickness parasite Trypanosoma brucei was solved by molecular replacement at 3.2-A resolution with an x-ray data set collected by the Laue method. For data collection, three crystals were exposed to the polychromatic synchrotron x-ray beam for a total of 20.5 sec. The structure was solved by using the Bacillus stearothermophilus enzyme model [Skarzy?ski, T., Moody, P. C. E. & Wonacott, A. J. (1987) J. Mol. Biol. 193, 171-187] with a partial data set which was 37% complete. The crystals contain six subunits per asymmetric unit, which allowed us to overcome the absence of > 60% of the reflections by 6-fold density averaging. After molecular dynamics refinement, the current molecular model has an R factor of 17.6%. Comparing the structure of the trypanosome enzyme with that of the homologous human muscle enzyme, which was determined at 2.4-A resolution, reveals important structural differences in the NAD binding region. These are of great interest for the design of specific inhibitors of the parasite enzyme.

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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC46085PMC
http://dx.doi.org/10.1073/pnas.90.6.2355DOI Listing
March 1993
78 Reads
107 Citations
9.809 Impact Factor

Three-dimensional structure of the quinoprotein methylamine dehydrogenase from Paracoccus denitrificans determined by molecular replacement at 2.8 A resolution.

Proteins 1992 Oct;14(2):288-99

Department of Cell Biology and Physiology, Washington University School of Medicine, St. Louis, Missouri 63110.

The three-dimensional structure of the quinoprotein methylamine dehydrogenase from Paracoccus dentrificans (PD-MADH) has been determined at 2.8 A resolution by the molecular replacement method combined with map averaging procedures, using data collected from an area detector. The structure of methylamine dehydrogenase from Thio-bacillus versutus, which contains an "X-ray" sequence, was used as the starting search model. MADH consists of 2 heavy (H) and 2 light (L) subunits related by a molecular 2-fold axis. The H subunit is folded into seven four-stranded beta segments, forming a disk-shaped structure, arranged with pseudo-7-fold symmetry. A 31-residue elongated tail exists at the N-terminus of the H subunit in MADH from T. versutus but is partially digested in this crystal form of MADH from P. denitrificans, leaving the H subunit about 18 residues shorter. Each L subunit contains 127 residues arranged into 10 beta-strands connected by turns. The active site of the enzyme is located in the L subunit and is accessible via a hydrophobic channel between the H and L subunits. The redox cofactor of MADH, tryptophan tryptophylquinone is highly unusual. It is formed from two covalently linked tryptophan side chains at positions 57 and 107 of the L subunit, one of which contains an orthoquinone.

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http://dx.doi.org/10.1002/prot.340140214DOI Listing
October 1992
4 Reads
60 Citations
2.627 Impact Factor

Crystal structure of an electron-transfer complex between methylamine dehydrogenase and amicyanin.

Biochemistry 1992 Jun;31(21):4959-64

Department of Cell Biology and Physiology, Washington University School of Medicine, St. Louis, Missouri 63110.

The crystal structure of the complex between the quinoprotein methylamine dehydrogenase (MADH) and the type I blue copper protein amicyanin, both from Paracoccus denitrificans, has been determined at 2.5-A resolution using molecular replacement. The search model was MADH from Thiobacillus versutus. The amicyanin could be located in an averaged electron density difference map and the model improved by refinement and model building procedures. Nine beta-strands are observed within the amicyanin molecule. The copper atom is located between three antiparallel strands and is about 2.5 A below the protein surface. The major intermolecular interactions occur between amicyanin and the light subunit of MADH where the interface is largely hydrophobic. The copper atom of amicyanin and the redox cofactor of MADH are about 9.4 A apart. One of the copper ligands, His 95, lies between the two redox centers and may facilitate electron transfer between them.

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http://dx.doi.org/10.1021/bi00136a006DOI Listing
June 1992
5 Reads
197 Citations
3.015 Impact Factor

Crystallographic investigations of the tryptophan-derived cofactor in the quinoprotein methylamine dehydrogenase.

FEBS Lett 1991 Aug;287(1-2):163-6

Department of Cell Biology and Physiology, Washington University School of Medicine, St. Louis, MO 63110.

A model of tryptophan tryptophylquinone (TTQ), recently proposed by McIntire et al. (Science (1991) 252, 817-824) to be the prosthetic group of the quinoprotein methylamine dehydrogenase, has been compared with electron density maps of this dehydrogenase from Thiobacillus versutus and Paracoccus denitrificans. The comparison shows that the TTQ model can be neatly accommodated, providing strong supportive evidence that TTQ is indeed the cofactor for this group of quinoproteins.

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http://dx.doi.org/10.1016/0014-5793(91)80041-zDOI Listing
August 1991
4 Reads
72 Citations
3.169 Impact Factor

The cytosolic and glycosomal glyceraldehyde-3-phosphate dehydrogenase from Trypanosoma brucei. Kinetic properties and comparison with homologous enzymes.

Eur J Biochem 1991 Jun;198(2):429-35

International Institute of Cellular and Molecular Pathology, Research Unit for Tropical Diseases, Brussels, Belgium.

The protozoan haemoflagellate Trypanosoma brucei has two NAD-dependent glyceraldehyde-3-phosphate dehydrogenase isoenzymes, each with a different localization within the cell. One isoenzyme is found in the cytosol, as in other eukaryotes, while the other is found in the glycosome, a microbody-like organelle that fulfils an essential role in glycolysis. The kinetic properties of the purified glycosomal and cytosolic isoenzymes were compared with homologous enzymes from other organisms. Both trypanosome enzymes had pH/activity profiles similar to that of other glyceraldehyde-3-phosphate dehydrogenases, with optimal activity around pH 8.5-9. Only the yeast enzyme showed its maximal activity at a lower pH. The glycosomal enzyme was more sensitive to changes in ionic strength below 0.1 M, while the cytosolic enzyme resembled more the enzymes from rabbit muscle, human erythrocytes and yeast. The affinity for NAD of the glycosomal enzyme was 5-10-fold lower than that of the cytosolic, as well as the other enzymes. A similar, but less pronounced, difference was found for its affinity for NADH. These differences are explained by a number of amino acid substitutions in the NAD-binding domain of the glycosomal isoenzyme. In addition, the effects of suramin, gossypol, agaricic acid and pentalenolactone on the trypanosome enzymes were studied. The trypanocidal drug suramin inhibited both enzymes, but in a different manner. Inhibition of the cytosolic enzyme was competitive with NAD, while in the case of the glycosomal isoenzyme, with NAD as substrate, the drug had an effect both on Km and Vmax. The most potent inhibitor was pentalenolactone, which at micromolar concentrations inhibited the glycosomal enzyme and the enzymes from yeast and Bacillus stearothermophilus in a reversible manner, while the rabbit muscle enzyme was irreversibly inhibited.

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http://dx.doi.org/10.1111/j.1432-1033.1991.tb16032.xDOI Listing
June 1991
3 Reads
91 Citations
4.530 Impact Factor

Structure determination of quinoprotein methylamine dehydrogenase from Thiobacillus versutus.

Acta Crystallogr B 1990 Dec;46 ( Pt 6):806-23

Department of Chemistry, University of Groningen, The Netherlands.

The crystal structure of quinoprotein methylamine dehydrogenase from Thiobacillus versutus (EC 1.4.99.3, Mr = 123,500) has been solved to 2.25 A resolution. The crystals of space group P3(1)21 (a = b = 129.8, c = 104.3 A) contain half a tetrameric enzyme molecule in the asymmetric unit, with a solvent content of ca 70%. The procedure used to solve this structure involved multiple isomorphous-replacement phasing, complemented by phase extension using solvent flattening, and phase combination with partial-model phases. The use of solvent flattening was essential to generate good quality electron density maps into which initial models were built. These partial models were refined using molecular-dynamics procedures. Refined model phases were then combined with solvent-flattening phases to generate improved electron density distributions. In the absence of an amino-acid sequence for this enzyme, the current 2.25 A resolution electron density map was interpreted to provide a model for the complete molecule. The crystallographic R factor for this model, which lacks any water molecules, is 28.6% for data between 6.0 and 2.25 A resolution.

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http://dx.doi.org/10.1107/s010876819000636xDOI Listing
December 1990
2 Reads
15 Citations
2.184 Impact Factor

Three-dimensional structure of quinoprotein methylamine dehydrogenase

Authors:
FM Vellieux

Proefschrift Rijsuniversiteit Groningen 1990

Rijksuniversiteit Groningen

Proefschrift ter verkrijging van het doktoraat in de Wiskunde en Natuurwetenschappen aan de Rijksuniversiteit Groningen

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June 1990

1 Citation

A new model for the pro-PQQ cofactor of quinoprotein methylamine dehydrogenase

FEBS Lett. 1989 Sep 25:255(2):460

FEBS Lett.

AbstractA model for the pro-PQQ cofactor of Thiobucillus versutus methylamine dehydrogenase was fitted into a 2.25 A resolution electron density distribution for this enzyme. This proposed model of pro-PQQ consists of a tyrosine-derived quinone indole bicyclic structure. Linkage of the cofactor to the light subunit of the enzyme occurs via the side chain of glutamate 57, which is itself bound to the side chain of arginine 107 of that subunit. Since the elements of this cofactor are derived from tyrosine, glutamate and arginine, we have named it ‘TGA pro-PQQ’.

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September 1989

18 Citations

Impact Factor 2.999

Structure of quinoprotein methylamine dehydrogenase at 2.25 A resolution.

EMBO J 1989 Aug;8(8):2171-8

Laboratory of Chemical Physics, University of Groningen, Nijenborgh, The Netherlands.

The three-dimensional structure of quinoprotein methylamine dehydrogenase from Thiobacillus versutus has been determined at 2.25 A resolution by a combination of multiple isomorphous replacement, phase extension by solvent flattening and partial structure phasing using molecular dynamics refinement. In the resulting map, the polypeptide chain for both subunits could be followed and an X-ray sequence was established. The tetrameric enzyme, made up of two heavy (H) and two light (L) subunits, is a flat parallellepiped with overall dimensions of approximately 76 x 61 x 45 A. The H subunit, comprising 370 residues, is made up of two distinct segments: the first 31 residues form an extension which embraces one of the L subunits; the remaining residues are found in a disc-shaped domain. This domain is formed by a circular arrangement of seven topologically identical four-stranded antiparallel beta-sheets, with approximately 7-fold symmetry. In spite of distinct differences, this arrangement is reminiscent of the structure found in influenza virus neuraminidase. The L subunit consists of 121 residues, out of which 53 form a beta-sheet scaffold of a central three-stranded antiparallel sheet flanked by two shorter two-stranded antiparallel sheets. The remaining residues are found in segments of irregular structure. This subunit is stabilized by six disulphide bridges, plus two covalent bridges involving the quinone co-factor and residues 57 and 107 of this subunit. The active site is located in a channel at the interface region between the H and L subunits, and the electron density in this part of the molecule suggests that the co-factor of this enzyme is not pyrrolo quinoline quinone (PQQ) itself, but might be instead a precursor of PQQ.

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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC401144PMC
August 1989
78 Reads
130 Citations
10.434 Impact Factor

Protein crystallography, computer graphics, and sleeping sickness

Mol. Recog.: Chem. & Biochem. Problems 1989 Apr 1:84

Molecular Recognition: Chemical and Biochemical Problems

Introduction: Towards more rational drug design

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April 1989

2 Citations

The use of solvent-flattening procedures in the crystal structure determination of quinoprotein methylamine dehydrogenase

Proc CCP4 Study Weekend 1988:88

Proceedings of the 1988 CCP4 Study Weekend

Quinoproteins are enzymes which contain the recently discovered cofactor pyrrolo-quinoline quinone, or PQQ.

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January 1989

Protein structure refinement by molecular dynamics techniques

Proc. CCP4 Study Weekend 1989:1

Proceedings of the 1989 CCP4 Study Weekend

Since the early seventies protein crystallographers have been trying to obtain a model of the protein under investigation agreeing as closely as possible with the observed data.

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January 1989

1 Citation

Purification, crystallization and preliminary X-ray investigation of quinoprotein methylamine dehydrogenase from Thiobacillus versutus.

Eur J Biochem 1986 Jan;154(2):383-6

The enzyme methylamine dehydrogenase or primary-amine:(acceptor) oxidoreductase (deaminating) (EC 1.4.99.3) was purified from the bacterium Thiobacillus versutus to homogeneity, as judged by polyacrylamide gel electrophoresis. The native enzyme has a Mr of 123 500 and contains four subunits arranged in a alpha 2 beta 2 configuration, the light and heavy subunits having a Mr of 12900 and 47500 respectively. The isoelectric point is 3.9. The purified enzyme was crystallized from 37--42% saturated ammonium sulphate in 0.1 M sodium acetate buffer, pH 5.0. The space group is P3(1)21 or P3(2)21, with one alpha 2 beta 2 molecule in the asymmetric unit. The cell dimensions are: a = b = 13.01 nm; c = 10.40 nm. The X-ray diffraction pattern extends to at least 0.25-nm resolution.

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http://dx.doi.org/10.1111/j.1432-1033.1986.tb09409.xDOI Listing
January 1986
3 Reads
44 Citations
4.530 Impact Factor