References of "Ghuysen, Jean-Marie"
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See detailCloning, sequencing and overexpression in Escherichia coli of the alginatelyase-encoding aly gene of Pseudomonas alginovora: identification of three classes of alginate lyases
Chavagnat, Frederic; Duez, Colette ULg; Guinand, Micheline et al

in Biochemical Journal (1996), 319(Pt 2), 575-583

A gene of Pseudomonas alginovora, called aly, has been cloned in Escherichia coli using a battery of PCR techniques and sequenced. It encodes a 210-amino-acid alginate lyase (EC 4.2.2.3), Aly, in the form ... [more ▼]

A gene of Pseudomonas alginovora, called aly, has been cloned in Escherichia coli using a battery of PCR techniques and sequenced. It encodes a 210-amino-acid alginate lyase (EC 4.2.2.3), Aly, in the form of a 233-amino-acid precursor. P. alginovora Aly has been overproduced in E. coli with a His-tag sequence fused at the C-terminal end under conditions in which the formation of inclusion bodies is avoided. His-tagged P. alginovora Aly has the same enzymic properties as the wild-type enzyme and has the specificity of a mannuronate lyase. It can be purified in a one-step procedure by affinity chromatography on Ni(2+)-nitriloacetate resin. The yield is of 5 mg of enzyme per litre of culture. The amplification factor is 12.5 compared with the level of production by wild-type P. alginovora. The six alginate lyases of known primary structure fall into three distinct classes, one of which comprises the pair P. alginovora Aly and Klebsiella pneumoniae Aly. [less ▲]

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See detailOverproduction and properties of the mannuronate alginate lyase AlxMB
Malissard, Martine; Chavagnat, Frédéric; Duez, Colette ULg et al

in FEMS Microbiology Letters (1995), 126(2), 105-111

In previous studies (Malissard et al., FEMS Microbiol. Lett. (1993) 110, 101-106), the alginate lyase AlxM of the marine bacterium ATCC 433367 was produced in Escherichia coli TC4/pAL-A3 with a yield of ... [more ▼]

In previous studies (Malissard et al., FEMS Microbiol. Lett. (1993) 110, 101-106), the alginate lyase AlxM of the marine bacterium ATCC 433367 was produced in Escherichia coli TC4/pAL-A3 with a yield of 50 mu g per litre of culture. The polypeptide chain was cleaved between two cysteine residues, C169 and C183, themselves linked by a disulphide bridge. AlxM has now been overproduced in E. coli BL21(DE3)/pAL-Sur/pLysS. Under conditions in which formation of inclusion bodies can be avoided, the enzyme is synthesized as a catalytically active, water-soluble, unnicked polypeptide with a yield of 32 mg per litre of culture. It has been purified to protein homogeneity using a one-step procedure. The nicked ALxM(A) and unnicked ALxM(B) alginate lyases have identical alginate-degrading activities at high salt concentrations. [less ▲]

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See detailStreptomyces K15 active-site serine DD-transpeptidase: specificity profile for peptide, thiol ester and ester carbonyl donors and pathways of the transfer reactions.
Grandchamps, Jacqueline; Distèche, Martine ULg; Damblon, Christian ULg et al

in Biochemical Journal (1995), 307(Pt 2), 335-339

The Streptomyces K15 transferase is a penicillin-binding protein presumed to be involved in bacterial wall peptidoglycan crosslinking. It catalyses cleavage of the peptide, thiol ester or ester bond of ... [more ▼]

The Streptomyces K15 transferase is a penicillin-binding protein presumed to be involved in bacterial wall peptidoglycan crosslinking. It catalyses cleavage of the peptide, thiol ester or ester bond of carbonyl donors Z-R1-CONH-CHR2-COX-CHR3-COO- (where X is NH, S or O) and transfers the electrophilic group Z-R1-CONH-CHR2-CO to amino acceptors via an acyl-enzyme intermediate. Kinetic data suggest that the amino acceptor behaves as a simple alternative nucleophile at the level of the acyl-enzyme in the case of thiol ester and ester donors, and that it binds to the enzyme.carbonyl donor Michaelis complex and influences the rate of enzyme acylation by the carbonyl donor in the case of amide donors. Depending on the nature of the scissile bond, the enzyme has different requirements for substituents at positions R1, R2 and R3. [less ▲]

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See detailMolecular Structures of Penicillin-Binding Proteins and Beta-Lactamases
Ghuysen, Jean-Marie ULg

in Trends in Microbiology (1994), 2(10), 372-380

In the past, new antibacterial agents have been selected either from natural sources or by 'trial and error' modification of existing antibacterials. Future therapeutic strategies are likely to depend on ... [more ▼]

In the past, new antibacterial agents have been selected either from natural sources or by 'trial and error' modification of existing antibacterials. Future therapeutic strategies are likely to depend on increased knowledge of existing drug targets and the search for new targets. The machinery for the assembly of bacterial-cell-wall peptidoglycan is an ideal place to look. [less ▲]

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See detailErratum for : primary structure of the streptomyces R61 extracellular DD-peptidase. 1. Cloning into Streptomyces lividans and nucleotide sequence of the gene
Duez, Colette ULg; Piron-Fraipont, C.; Joris, Bernard ULg et al

in European Journal of Biochemistry (1994), 224(3), 1079

This is the correction of the fig. 5 of Primary structure of the Streptomyces R61 extracellular DD-peptidase. 1. Cloning into Streptomyces Zividuns and nucleotide sequence of the gene, by C. Duez, C ... [more ▼]

This is the correction of the fig. 5 of Primary structure of the Streptomyces R61 extracellular DD-peptidase. 1. Cloning into Streptomyces Zividuns and nucleotide sequence of the gene, by C. Duez, C. Piron-Fraipont, B. Joris, J. Dusart, M. S. Urdea, J. A. Martial, J.-M. Frère and J.-M. Ghuysen. European Journal of Biochemistry Volume 162, Issue 3, pages 509–518, February 1987 [less ▲]

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See detailThe Precursor of the Streptomyces R61 Dd-Peptidase Containing a C-Terminal Extension Is Inactive
Fanuel, Laurence; Granier, Benoît; Wilkin, Jean-Marc et al

in FEBS Letters (1994), 351(1), 49-52

The Streptomyces R61 DD-peptidase gene encodes a 26-residue C-terminal extension which is not found in the mature protein. When the gene was expressed in Escherichia coli, the extension was not cleaved ... [more ▼]

The Streptomyces R61 DD-peptidase gene encodes a 26-residue C-terminal extension which is not found in the mature protein. When the gene was expressed in Escherichia coli, the extension was not cleaved and the precursor protein was not enzymatically active. It also reacted with penicillins significantly more slowly than the mature protein. The introduction of a 'stop' codon after that corresponding to the C-terminal residue of the mature protein resulted in the production of an active protein in the periplasm of E. coli. [less ▲]

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See detailSite-directed mutagenesis of penicillin-binding protein 3 of Escherichia coli: role of Val-545
Ayala, Juan; Goffin, Colette; Nguyen-Distèche, Martine ULg et al

in FEMS Microbiology Letters (1994), 121(2), 251-256

Val545 of the Escherichia coli penicillin-binding protein 3 is essential to the acyl transfer mechanism through which the active-site serine 307 is acylated by benzylpenicillin and cephalexin and to the ... [more ▼]

Val545 of the Escherichia coli penicillin-binding protein 3 is essential to the acyl transfer mechanism through which the active-site serine 307 is acylated by benzylpenicillin and cephalexin and to the mechanism through which the protein allows rapidly growing cells to divide. [less ▲]

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See detailBinding Site-Shaped Repeated Sequences of Bacterial Wall Peptidoglycan Hydrolases
Ghuysen, Jean-Marie ULg; Lamotte-Brasseur, Josette; Joris, Bernard ULg et al

in FEBS Letters (1994), 342(1), 23-28

The non-catalytic C-terminal regions of the N-acetylmuramidase (lysozyme) of Clostridium acetobutylicum and N-acetylmuramoyl(D-lactyl)-L-alanine amidases CwlA of Bacillus subtilis, ORFL3 and CwlL of ... [more ▼]

The non-catalytic C-terminal regions of the N-acetylmuramidase (lysozyme) of Clostridium acetobutylicum and N-acetylmuramoyl(D-lactyl)-L-alanine amidases CwlA of Bacillus subtilis, ORFL3 and CwlL of Bacillus licheniformis were previously reported to have similarities with the amino acid sequence of the non-catalytic N-terminal module of the Streptomyces albus G Zn DD-peptidase. This peptidase is a bipartite protein of known three-dimensional structure. Its non-catalytic N-terminal module possesses, exposed at the surface, an elongated crevice which is defined by a loop-helix-loop-helix motif that consists of two repeats, each 16 amino acid residues long, connected by a heptapeptide and whose design is compatible with its possible functioning as a substrate recognition and binding site. Amino acid alignments suggest that cavities nearly identical in shape to that present in the non-catalytic module of the S. albus peptidase, are borne by the C-terminal regions of the CwlA amidase (in one copy), the lysozyme and the ORFL3 and CwlL amidases (in two copies). Since a common feature of the five enzymes is their substrate, the bacterial cell wall peptidoglycan, we interpret the striking similarity of their non-catalytic N- or C-terminal modules to suggest that these modules are involved in the binding of these exocellular enzymes to their insoluble wall substrate. [less ▲]

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See detailEngineering and Overexpression of Periplasmic Forms of the Penicillin-Binding Protein 3 of Escherichia Coli
Fraipont, Claudine ULg; Adam, Maggy; Nguyen-Disteche, Martine et al

in Biochemical Journal (1994), 298(1), 189-195

Replacement of the 36 and 56 N-terminal amino acid residues of the 588-amino-acid-residue membrane-bound penicillin-binding protein 3 (PBP3) of Escherichia coli by the OmpA signal peptide allows export of ... [more ▼]

Replacement of the 36 and 56 N-terminal amino acid residues of the 588-amino-acid-residue membrane-bound penicillin-binding protein 3 (PBP3) of Escherichia coli by the OmpA signal peptide allows export of F37-V577 PBP3 and G57-V577 PBP3 respectively into the periplasm. The modified ftsI genes were placed under the control of the fused lpp promoter and lac promoter/operator; expression of the truncated PBP3s was optimized by varying the copy number of the recombinant plasmids and the amount of LacI repressor, and export was facilitated by increasing the SecB content of the producing strain. The periplasmic PBP3s (yield 8 mg/l of culture) were purified to 70% protein homogeneity. They require the presence of 0.25 M NaCl to remain soluble. Like the membrane-bound PBP3, they undergo processing by elimination of the C-terminal decapeptide I578-S588, they bind penicillin in a 1:1 molar ratio and they catalyse hydrolysis and aminolysis of acyclic thioesters that are analogues of penicillin. The membrane-anchor-free PBP3s have ragged N-termini. The G57-V577 PBP3, however, is less prone to proteolytic degradation than the F37-V577 PBP3. [less ▲]

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See detailBacterial cell wall
Ghuysen, Jean-Marie ULg; Hackenbeck, Regine

Book published by Elsevier (1994)

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See detailBiochemistry of the penicilloyl serine transferases
Ghuysen, Jean-Marie ULg; Dive, Georges ULg

in Ghuysen, Jean-Marie; Hackenbeck, Regine (Eds.) Bacterial cell wall (1994)

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See detailSerine-Type D-Ala-D-Ala Peptidases and Penicillin-Binding Proteins
Granier, Benoît; Jamin, Marc; Adam, Maggy et al

in Methods in Enzymology (1994), 244

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See detailDetailed Study of a Molecule in a Molecule N-Acetyl-L-Tryptophanamide in a Active-Site Model of Alpha-Chymotrypsin
Dive, Georges ULg; Dehareng, Dominique ULg; Ghuysen, Jean-Marie ULg

in Journal of the American Chemical Society (1994), 116(6), 2548-2556

Six complexes between a model active site of alpha-chymotrypsin (261 atoms) and N-acetyl-L-tryptophanamide (33 atoms) were optimized at the semiempirical AM1 level. In one of these complexes, a water ... [more ▼]

Six complexes between a model active site of alpha-chymotrypsin (261 atoms) and N-acetyl-L-tryptophanamide (33 atoms) were optimized at the semiempirical AM1 level. In one of these complexes, a water molecule was included. A detailed study at the geometric and energetic levels is presented. The discussion deals with the nature of the interaction, the effect of the environment, the ligand deformation, the backbone relaxation, the water molecule freedom, and the comparison with the molecular mechanics results. [less ▲]

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See detailSite-directed mutagenesis of dicarboxylic acid residues of the penicillin-binding module of the Escherichia coli penicillin-binding protein 3
Goffin, Colette ULg; Ayala, J. A.; Nguyen-Distèche, Martine ULg et al

in FEMS Microbiology Letters (1993), 113(3), 247-251

The glutamic acid E396, aspartic acid D409 and glutamic acid E411 residues of the Escherichia coli penicillin-binding protein 3 were each converted into an alanine residue. As deduced from penicillin ... [more ▼]

The glutamic acid E396, aspartic acid D409 and glutamic acid E411 residues of the Escherichia coli penicillin-binding protein 3 were each converted into an alanine residue. As deduced from penicillin-binding and complementation experiments, none of these dicarboxylic acid residues is involved in the mechanism of acylation by penicillin and none of them is essential for the in vivo functioning of the PBP. The mutation E396, however, causes an increased thermolability of the protein. [less ▲]

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See detailModular Design of the Bi(multi) functional penicillin-binding proteins
Englebert, Serge; El Kharroubi, Aboubaker; Piras, Graziella et al

in De pedro; Höltje, J. V.; Loffelhardt, W. (Eds.) Bacterial Growth & Lysis Metabolism and Structure of the Bacterial Sacculus (1993, June 30)

Proceedings of a symposium held in Mallorca, Spain in April 1992. The goal of the meeting was to assess the present state of knowledge on the structure and physiology of the bacterium murien sacculus, and ... [more ▼]

Proceedings of a symposium held in Mallorca, Spain in April 1992. The goal of the meeting was to assess the present state of knowledge on the structure and physiology of the bacterium murien sacculus, and develop new hypotheses and strategies to promote further development of the field. The contributions reflect broadly different approaches. Papers discuss structure and chemistry, biosynthesis and maturation, regulation and control of cell wall hydrolases, penicillin interactive proteins, morphogenesis and septum formation, and cell growth. Annotation c. Book News, Inc., Portland, OR (booknews.com) [less ▲]

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See detailCharacterization of the Sporulation-Related Gamma-D-Glutamyl-(L)Meso-Diaminopimelic-Acid-Hydrolysing Peptidase I of Bacillus Sphaericus NCTC 9602 as a Member of the Metallo(Zinc) Carboxypeptidase A Family. Modular Design of the Protein
Hourdou, Marie-Laure; Guinand, Micheline; Vacheron, Marie-Jeanne et al

in Biochemical Journal (1993), 292(Pt 2), 563-570

The sporulation-related gamma-D-glutamyl-(L)meso-diaminopimelic-acid-hydrolysing peptidase I of Bacillus sphaericus NCTC 9602 has been analysed by proton-induced X-ray emission. It contains 1 equivalent ... [more ▼]

The sporulation-related gamma-D-glutamyl-(L)meso-diaminopimelic-acid-hydrolysing peptidase I of Bacillus sphaericus NCTC 9602 has been analysed by proton-induced X-ray emission. It contains 1 equivalent Zn2+ per mol of protein. As derived from gene cloning and sequencing, the B. sphaericus Zn peptidase I is a two-module protein. A 100-amino-acid-residue N-terminal domain consisting of two tandem segments of similar sequences, is fused to a 296-amino-acid-residue C-terminal catalytic domain. The catalytic domain belongs to the Zn carboxypeptidase A family, the closest match being observed with the Streptomyces griseus carboxypeptidase [Narahashi (1990) J. Biochem. 107, 879-886] and with the family prototype, bovine carboxypeptidase A. The catalytic domain of the B. sphaericus peptidase I possesses, distributed along the amino-acid sequence, peptide segments, a triad His162-Glu165-His307 and a dyad Tyr347-Glu366 that are equivalent to secondary structures, the zinc-binding triad His69-Glu72-His196 and the catalytic dyad Tyr248-Glu270 of bovine carboxypeptidase A respectively. The N-terminal repeats of the B. sphaericus peptidase I have similarity with the C-terminal repeats of the Enterococcus hirae muramidase 2, the Streptococcus (now Enterococcus) faecalis autolysin and the Bacillus phi PZA and phi 29 lysozymes, to which a role in the recognition of a particular moiety of the bacterial cell envelope has been tentatively assigned. Detergents enhance considerably the specific activity of the B. sphaericus peptidase I. [less ▲]

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See detailCloning and Sequencing of the Low-Affinity Penicillin-Binding Protein 3r-Encoding Gene of Enterococcus Hirae S185: Modular Design and Structural Organization of the Protein
Piras, Graziella; Raze, Dominique; el Kharroubi, Aboubaker et al

in Journal of Bacteriology (1993), 175(10), 2844-2852

The clinical isolate Enterococcus hirae S185 has a peculiar mode of resistance to penicillin in that it possesses two low-affinity penicillin-binding proteins (PBPs): the 71-kDa PBP5, also found in other ... [more ▼]

The clinical isolate Enterococcus hirae S185 has a peculiar mode of resistance to penicillin in that it possesses two low-affinity penicillin-binding proteins (PBPs): the 71-kDa PBP5, also found in other enterococci, and the 77-kDa PBP3r. The two PBPs have the same low affinity for the drug and are immunochemically related to each other. The PBP3r-encoding gene has been cloned and sequenced, and the derived amino acid sequence has been compared by computer-assisted hydrophobic cluster analysis with that of the low-affinity PBP5 of E. hirae R40, the low-affinity PBP2' of Staphylococcus aureus, and the PBP2 of Escherichia coli used as the standard of reference of the high-M(r) PBPs of class B. On the basis of the shapes, sizes, and distributions of the hydrophobic and nonhydrophobic clusters along the sequences and the linear amino acid alignments derived from this analysis, the dyad PBP3r-PBP5 has an identity index of 78.5%, the triad PBP3r-PBP5-PBP2' has an identity index of 29%, and the tetrad PBP3r-PBP5-PBP2'-PBP2 (of E. coli) has an identity index of 13%. In spite of this divergence, the low-affinity PBPs are of identical modular design and possess the nine amino acid groupings (boxes) typical of the N-terminal and C-terminal domains of the high-M(r) PBPs of class B. At variance with the latter PBPs, however, the low-affinity PBPs have an additional approximately 110-amino-acid polypeptide stretch that is inserted between the amino end of the N-terminal domain and the carboxy end of the membrane anchor. While the enterococcal PBP5 gene is chromosome borne, the PBP3r gene appears to be physically linked to the erm gene, which confers resistance to erythromycin and is known to be plasmid borne in almost all the Streptococcus spp. examined. [less ▲]

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See detailEnergy Analysis on Small to Medium-Sized H-Bonded Complexes
Dive, Georges ULg; Dehareng, Dominique ULg; Ghuysen, Jean-Marie ULg

in Theoretica Chimica Acta (1993), 85(6), 409-421

Dimers (water-methanol, guanine-cytosine) as well as trimers (methanol-water-imidazole, formamide-methylformate-formamide), are studied as H-bonded complexes of increasing complexity. All the investigated ... [more ▼]

Dimers (water-methanol, guanine-cytosine) as well as trimers (methanol-water-imidazole, formamide-methylformate-formamide), are studied as H-bonded complexes of increasing complexity. All the investigated conformations have been fully optimized. In particular, it is the first time that all the intra- and intermolecular parameters of the guanine-cytosine complex are left variable. In minimal basis sets, the planar conformation has been found to be a first-order critical point. The minimal basis set MINI-1 has been adapted to provide nearly planar amides. The stability of the complexes is accounted for by four energy components of the same order: the first-order term (electrostatic + exchange), the polarization, the charge transfer and the correlation terms. In the case of the studied trimers, the energy components, apart from the electrostatic one, have been found to be nearly additive. [less ▲]

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See detailAnalytical calculation of the electrostatic interaction energy within the CNDO framework
Dehareng, Dominique ULg; Dive, Georges ULg; Ghuysen, Jean-Marie ULg

in International Journal of Quantum Chemistry (1993), 46(6), 711-734

This work analyses the adequacy of an analytical electrostatic energy formulation within the CNDO framework to predict the stable conformations of large molecular complexes. Comparisons are made with ab ... [more ▼]

This work analyses the adequacy of an analytical electrostatic energy formulation within the CNDO framework to predict the stable conformations of large molecular complexes. Comparisons are made with ab initio results for small systems such as water-formamide, methanol-water-imidazole, or guanine-cytosine and with AM1 results for two large systems: a molecular tweezer + the 9-methyladenine complex and a model active site of the alpha-chymotrypsin and its ligand complex. This approach is efficient in providing reliable conformers for large molecular systems in a very fast way. [less ▲]

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See detailPolarization-corrected electrostatic potentials of aromatic compounds
Dehareng, Dominique ULg; Dive, Georges ULg; Ghuysen, Jean-Marie ULg

in Journal of the American Chemical Society (1993), 115(15), 6877-6882

The electrostatic potentials (EPS) corrected for polarization (TPS) of the aromatic compounds benzene, aniline, chlorobenzene, nitrobenzene, phenol, benzamide, and N-phenylacetamide have been calculated ... [more ▼]

The electrostatic potentials (EPS) corrected for polarization (TPS) of the aromatic compounds benzene, aniline, chlorobenzene, nitrobenzene, phenol, benzamide, and N-phenylacetamide have been calculated at the ab initio SCF level within three basis sets: 6-31G**, MINI-1, and STO-3G. For chlorobenzene in its MINI-1-optimized geometry, the calculation was also performed within MINI-1**. By reference to 6-31G**, the MINI-1-computed EP is much more satisfactory than the STO-3G-computed EP, whereas the MINI-I and STO-3G basis sets give very similar total potentials corrected for polarization (TPs). The MINI-1** basis set appears to be miscalibrated for computing EPs. It provides qualitative results that differ from those obtained with the 6-31G** basis set. The EP has a negative well above the middle of the benzene ring, while the TP exhibits a negative crown just above the benzene carbon atoms, where electrophilic attack takes place. The TP calculated for the interaction of nitrobenzene with a hydride ion instead of a proton allowed analyzation of the effects of polarization on the positive EP above the N-C bond. [less ▲]

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